Study on the mechanism of chiral recognition with molecularly imprinted polymers

This study aimed at elucidating the chiral recognition mechanism with molecularly imprinted polymers (MIPs) in aqueous environment. The system used ethylene glycol dimethacrylate (EGDMA), methacrylic acid (MAA), and 4-l-phenylalanylamino-pyridine (4-l-PheNHPy) as the cross-linking monomer, functional monomer and template, respectively, to assemble the imprinted polymer. A self-assembly mechanism, which includes the pre-organizing functional monomers around template before polymerization process, was proposed. This mechanism was supported by NMR titration test. Interactions between functional monomer and template were observed using UV-Vis spectroscopy of solutions of these components as well. These studies indicated a 1:2 molecular complex dominantly formed between 4-l-PheNHPy and MAA. Association constant was estimated to be 97,000 M⁻². Based on these results, a model mainly involving two-spot interaction was proposed evolving from our reported concept of exact placement of functional group. Ionic interaction between the primary amino group of 4-l-PheNHPy and carboxylic acid group inside the microcavity on MIPs was believed to play a predominate role in the enantioselectivity as supported by the observation of the relationship between the retention factor of 4-l-PheNHPy and the pH of mobile phase. While thermodynamic study at different pH revealed that, the interaction between the pyridyl group of 4-l-PheNHPy and the carboxylic acid group on the MIPs is also strong, implying that it also plays a profound role in determining the highly chiral selectivity of MIPs.     

Monodispersed, molecularly imprinted polymers for cinchonidine by precipitation polymerization

Three monodispersed, molecularly imprinted polymers (MIPs) for cinchonidine (CD) have been synthesized by precipitation polymerization. MIP1 was prepared using methacrylic acid (MAA) as a functional monomer and divinylbenzene (DVB) as a cross-linker and MIP2 was prepared with further addition of 2-hydroxyethyl methacrylate (HEMA) as a co-monomer. For the preparation of MIP3, core-shell type MIP, monodispersed DVB homopolymers, which are prepared by precipitation polymerization, were used as a core and CD-imprinted MAA-DVB copolymer phases were coated onto the core. Three MIPs synthesized gave monodispersed, spherical beads in micrometer sizes. The binding characteristics and molecular recognition properties of MIP1-3 were examined by Scatchard analysis and chromatographic studies. The association constant of CD with MIP1 was the highest among MIPs prepared, while that with MIP3 was the lowest. The template molecule, CD, was more retained than its stereoisomer, cinchonine, on the three MIPs, and the stereoseparation factor of 38 was obtained with MIP3.     

Development and application of molecularly imprinted polymers as solid-phase sorbents for erythromycin extraction

Six molecularly imprinted polymers (MIPs) of erythromycin (ERY) were prepared by noncovalent bulk polymerization using methacrylic acid (MAA) as the functional monomer. On the basis of binding analysis, the MIPs with 1:2 optimum ratio of template to MAA were selected for subsequent scanning electron microscopy and Brunauer–Emmett–Teller analyses, which indicated that the MIPs had more convergent porous structures than the nonimprinted polymers. The equilibrium binding experiments showed that the binding sites of MIPs were heterogeneous, with two dissociation constants of 0.005 and 0.63 mg mL⁻¹, respectively. Furthermore, the performance of the MIPs as solid-phase extraction (SPE) sorbents was evaluated, and the selectivity analysis showed that the MIPs could recognize ERY with moderate cross-reactivity for other macrolides. The overall investigation of molecularly imprinted SPE for cleanup and enrichment of the ERY in pig muscle and tap water confirmed the feasibility of utilizing the MIPs obtained as specific SPE sorbents for ERY extraction in real samples. Figure Schematic diagram of the preparation and application of the erythromycin imprinted molecularly imprinted polymers Suquan Song and Aibo Wu contributed equally to this work.     

Improving the strategy and performance of molecularly imprinted polymers using cross-linking functional monomers

A new strategy for monomer design has been investigated that combines interactive monomer functionality with a cross-linking format, giving as a result noncovalent molecularly imprinted polymers (MIPs) with improved performance. This strategy was explored under the premise that more functionality could be introduced without suffering performance losses due to reduced cross-linking. While this proved to be correct, equally important contributions to selectivity enhancement at the molecular level by conformation control and diastereomeric complexation were also discovered. Monomers derived from l-serine and l-aspartic acid were synthesized and used to prepare MIPs, with the best performance obtained for the MIP formulated with the serine-based cross-linker (N,O-bis-methacryloyl l-serine, 3), versus the aspartic-acid-based cross-linkers and the traditional methacrylic acid/ethylene glycol dimethacrylate (MAA/EGDMA) formulation. Quantitative structure-selectivity relationship (QSSR) studies revealed that the improved performance of 3 was due to three key factors: (1) the cross-linking nature of this monomer; (2) control of conformational flexibility; (3) a strong influence of monomer chirality on enantioselectivity in MIPs.     

Clozapine imprinted polymers: synthesis, characterization and application for drug assay in human serum

A variety of molecularly imprinted polymers (MIPs) against clozapine (CLZ) were synthesized and their recognition properties were compared with blank non-imprinted polymers. Methacrylic acid (MAA) was used as a functional monomer and Chloroform or tetrahydrofuran (THF) were applied as polymerization solvents. Chloroform as the solvent and MAA/CLZ ratio of 5 was selected as optimized polymerization condition. In Scatchard analysis of MIP-CLZ interactions, two classes of binding sites were found in MIP—high affinity (KD = 14.5 μM) and low affinity (KD = 322.5 μM) binding sites. The polymer was evaluated as a selective sorbent in molecularly imprinted solid-phase extraction (MISPE) of CLZ from human serum. The MISPE procedure was developed and optimized with a recovery of 88-102%. The intra- and inter-day precision values were less than 1.36% and 2.5%, respectively. The selectivity of MISPE for CLZ was studied in comparison with some drugs. These drugs could be present with CLZ, simultaneously in serum of patients. The data indicated that the imprinted polymer had a good selectivity and affinity for CLZ and could be used for selective extraction and analysis of CLZ in human serum.     

Synthesis by precipitation polymerisation of molecularly imprinted polymer microspheres for the selective extraction of carbamazepine and oxcarbazepine from human urine

Two molecularly imprinted polymers (MIPs), in the physical form of well-defined polymer microspheres, were synthesised via precipitation polymerisation (PP) using an antiepileptic drug, carbamazepine (CBZ), as template molecule, methacrylic acid as functional monomer and either divinylbenzene 80 (DVB-80) or a mixture of DVB-80 and ethylene glycol dimethacrylate (EGDMA) as crosslinking agents. The MIP obtained using DVB-80 alone as crosslinking agent (MIP A) had a narrow particle size distribution (9.5 ± 0.5 μm) and a well-developed permanent pore structure (specific surface area in the dry state = 758 m² g⁻¹), whereas when a mixture of DVB-80 and EGDMA (MIP B) were used as crosslinking agents, the polymer obtained had a broader particle size distribution (6.4 ± 1.8 μm) and a relatively low specific surface area (23 m² g⁻¹). The molecular recognition character of both polymers was evaluated by means of LC and then a molecularly imprinted solid-phase extraction (MISPE) protocol; CBZ was recognised by both polymers, and useful cross-selectivity for oxcarbazepine (OCBZ), which is the main metabolite of CBZ, also observed. In a detailed bioanalytical study, MIP A was selected in preference to MIP B since MIP A enabled a high volume of sample to be extracted such that lower limits of detection were achievable using this polymer. High recoveries of CBZ and OCBZ were obtained in a MISPE protocol when 50 mL of human urine spiked at 0.2 mg L⁻¹ were percolated through MIP A (90% and 83%, respectively).     

Chiral recognition and separation of beta2-amino acids using non-covalently molecularly imprinted polymers

Non-covalently molecularly imprinted polymers (MIPs) for beta2-amino acids were prepared for the first time. N-(2-chlorobenzyloxycarbonyl)-(R)-beta2-homophenylalanine (N-2-ClZ-(R)-beta2-HPhe) was imprinted with methacrylic acid (MAA) and/or 4-vinylpyridine (4-VPy) as the functional monomers, with ethylene glycol dimethacrylate (EDMA) as the cross-linker. The MIPs made with different ratios of MAA:4-VPy were studied in HPLC mode. The results show that MIPs made with 4-VPy yielded the best chiral separation factor (alpha= 1.86) for the template molecule. The importance for an efficient separation of pi-stacking interactions between the MIPs and the template molecule is demonstrated. Racemates of Z-alpha-amino acids and beta-amino acid analogues of the template were either not or poorly resolved by the MIPs, thus demonstrating the close three-dimensional complementarity of the MIPs' recognition sites with the template.     

Chromatographic characterization of molecularly imprinted polymers

Recent efforts in the investigation of chromatographic characterization of molecularly imprinted polymers (MIPs) have focused mainly on the nature of heterogeneous binding sites. More data on the thermodynamics than on the kinetic features of MIP columns have been published. The present article addresses the sources of peak broadening and tailing, which are the main drawbacks often associated with imprinted polymers in chromatography for practical applications. With use of the theory of nonlinear chromatography, the peak properties of a MIP column, including the retention and peak broadening and tailing, can be well interpreted. Efforts to improve chromatographic efficiency using MIPs prepared by approaches different from the conventional method, including covalent imprinting and the format of uniformly sized spherical microbeads, are reviewed and discussed. This review leads to the conclusion that nonlinear chromatography theory is useful for characterizing chromatographic features of MIP columns, since a MIP is essentially an affinity-based chromatographic stationary phase. We expect more theoretical and experimental studies on the kinetic aspects of MIP columns, especially the factors influencing the apparent rate constant, as well as the analysis of the influences of mobile-phase composition on the chromatographic performance. In addition to revealing the affinity interaction by molecular recognition, slow nonspecific interactions which may be inherited from the imperfect imprinting and may be involved in the rebinding of the template to MIPs also need to be characterized. Figure The peak broadening and tailing associated often with molecularly imprinted polymers (MIPs) in column chromatography for practical applications can be well characterized by the theory of nonlinear chromatography.     

Selective solid-phase extraction of dibutyl phthalate from soybean milk using molecular imprinted polymers

An analysis method is reported for dibutyl phthalate and related compounds with high selectivity and sensitivity by using the selective molecularly imprinted solid-phase extraction (MISPE) technique. In this report, dibutyl phthalate (DBP) is employed as the template molecule, and the molecularly imprinted polymers (MIPs) are synthesized through the bulk polymerization of methacrylic acid (MAA). The Scatchard plot suggests that the template-polymer system has two-site binding behavior with the dissociation constants of 0.5187 and 0.01898 mmol L⁻¹, respectively. The rebinding test, based on the MISPE column technique, shows the recoveries of soybean milk samples spiked with 5 phthalates are in the range of 75.8-107.5% with the relative standard deviations of 1.80-10.08%, indicating the feasibility of the prepared MIPs for phthalates extraction. Finally, the method is used to analyze the trace level of phthalates in commercial soybean milk.     

Gas phase detection of explosives such as 2,4,6-trinitrotoluene by molecularly imprinted polymers

Fast, reliable and inexpensive analytical techniques for trace detection of explosive components are in high demand. Our approach is to develop specific sensor coating materials based on molecularly imprinted polymers (MIPs). Despite the known inhibition of radical polymerisations by nitro groups and the known shrinkage of the polymer lattice during/after drying we were able to synthesize particulate MIPs by suspension polymerisation as well as thin MIP coatings by direct surface polymerisation on quartz crystal microbalances (QCM). The best method to purify the porous beads was Soxhlet extraction followed by supercritical carbon dioxide extraction (SFE with sc-CO₂) at mild conditions (150 bar, 50 °C). At least a removal of >99.7% of the template was achieved. Performance tests of TNT imprinted polymer beads showed that acrylamide (AA) and more pronounced also methacrylic acid (MAA) possessed an enhanced adsorption tendency for gaseous TNT. An adsorption of 2,4-DNT, dinitrotoluene, by these MIPs was not detected. Using 2,4-DNT as template and methacrylamide, MAAM, a positive imprint effect for gaseous 2,4-DNT was achieved with no measurable cross-sensitivity for 2,4,6-TNT.The thin MIP coatings directly synthesized on the QCMs showed thicknesses of 20 to up to 500 nm. Preliminary screening experiments were performed for five different monomers and three different solvents (acetonitrile, chloroform and dimethylformamide). Best adsorption properties for TNT vapour until now showed a PAA-MIP synthesized with chloroform. Direct measurements of the mass attachment, respectively frequency decrease of the coated QCMs during vapour treatment showed a TNT-uptake of about 150 pg per μg MIP per hour. Results look worthy for further studies.     

Chiral resolution of derivatized amino acids using uniformly sized molecularly imprinted polymers in hydro-organic mobile phases

Uniformly sized molecularly imprinted polymers (MIPs) for Boc-l-Trp were prepared using ethylene glycol dimethacrylate (EDMA) as the cross-linker, and methacylic acid (MAA) and/or 4-vinylpyridine (4-VPY) as the functional monomers or without use of a functional monomer. The MIPs prepared were evaluated using acetonitrile or a mixture of phosphate buffer and acetonitrile as the mobile phase. The Boc-l-Trp-imprinted EDMA polymers can recognize Boc-l-Trp by its molecular shape, and can thus afford the enantioseparation of Boc-Trp. Besides the molecular shape recognition, the hydrophobic interactions with the polymer backbones as well as the hydrogen-bonding interactions of Boc-l-Trp with carboxyl and pyridyl groups in the polymers should work for the retention and recognition of Boc-l-Trp on the imprinted MAA-co-EDMA and 4-VPY-co-EDMA polymers, respectively, in the hydro-organic mobile phase. The hydrogen-bonding interactions seem to become dominant when only acetonitrile is used as the mobile phase. The Boc-l-Trp-imprinted 4-VPY-co-EDMA polymers gave the highest retentivity and enantioselectivity for Boc-Trp among the MIPs prepared. However, the simultaneous use of MAA and 4-VPY was not effective for the enantioseparation of Boc-Trp in a hydro-organic mobile phase. Furthermore, the baseline separation of Boc-Trp enantiomers was attained within 10 min on the Boc-l-Trp-imprinted 4-VPY-co-EDMA polymers under the optimized HPLC conditions.     

Computational prediction and experimental selectivity coefficients for hydroxyzine and cetirizine molecularly imprinted polymer based potentiometric sensors

In spite of the increasing usages number of molecularly imprinted polymers (MIPs) in many scientific applications, the theoretical aspects of participating intra molecular forces are not fully understood. This work investigates effects of the electrostatic force, the Mulliken charge and the role of cavity's backbone atoms on the selectivity of MIPs. Moreover, charge distribution, which is a computational parameter, was proposed for the prediction of the selectivity coefficients of MIP-based sensors. In the computational approaches and experimental study, methacrylic acid (MAA) was chosen as the functional monomer and ethylene glycol dimethacrylate (EGDMA) as the cross linker for hydroxyzine and cetirizine imprinted polymers. Ab initio, DFT B3LYP method was carried out on molecular optimization. With regard to results obtained from molecules optimization and hydrogen bonding properties, possible configurations of 1:n (n ≤ 5) template/monomer complexes were designed and optimized. The binding energy for each complex in gas phase was calculated. Depending on the most stable configuration, hydroxyzine and cetirizine imprinted polymer models were designed. The calculations including the porogen were also investigated. The theoretical charge distributions for the template and some potential interfering molecules were calculated. The results showed a correlation between the selectivity coefficients and the theoretical charge distributions. The results surprisingly show that charge distribution based model was able to predict the selectivity coefficients of MIP based potentiometric sensors.     

Molecularly imprinted polymers prepared in aqueous solution selective for [Sar,Ala]angiotensin II

Conventional molecular imprinting technology allows the synthesis in organic solvents of molecularly imprinted polymers (MIPs) selective toward relatively low molecular weight compounds. However, synthesis in aqueous media of chemically and mechanically stable MIPs that can recognize biomolecules such as peptides and proteins still is a great challenge. In this article, we report the successful synthesis of peptide-selective MIPs in aqueous solution. HPLC evaluation of these polymers with a water-based mobile phase showed their selectivity for the peptide, [Sar¹,Ala⁸]angiotensin II (SA), that had been used as the template, but not for its parent peptide angiotensin II (AII). The binding capacity and selectivity of our MIPs depended on the ratio of template to functional monomer in the polymerization mixture, as well as on the ionic strength and pH of the chromatographic mobile phase. These MIPs can be used for chromatographic detection of the octapeptide [Sar¹,Ala⁸]angiotensin II in aqueous solution, with a detection limit of 8 pmol and a response that is linear (r²>0.99) over the concentration range 0.4-20 μM.     

Novel stereoselective molecularly imprinted polymers via ring-opening metathesis polymerisation

Stereoselective molecularly imprinted polymers (MIPs) have been synthesised via ring-opening metathesis polymerisation, in essentially, quantitative yield. A covalent imprinting strategy was followed during the network formation of the chiral sorbent. Recognition of the substrate however involved non-covalent interactions; a combination of hydrogen bonding and the chiral environment presented by the imprinted cavities. The enantiomeric excess achievable with these new MIPs is solvent dependent and stereoselectivities of up to 20% e.e. (separation factor α=2.2) were found in batch equilibrations.     

Development of a group selective molecularly imprinted polymers based solid phase extraction of malachite green from fish water and fish feed samples

A group selective molecularly imprinted solid phase extraction (MISPE) for malachite green (MG) from fish water and fish feed samples was developed. Using MG as template molecule, methacrylic acid as functional monomer, ethylene glycoldimethacrylate as linking agent and bulk polymerization as synthetic method, the molecularly imprinted polymers (MIPs) were synthesized and characterized with rebinding experiment. The Scatchard polt's analysis revealed that the template-polymer system showed the two-site binding behavior with dissociation constants of 0.3194 μmol L⁻¹ and 15.70 μmol L⁻¹, respectively. MG and two structurally related compounds, leucomalachite green (LMG) and crystal violet (CV) were employed for selectivity test. The MIPs exhibited the highest selective rebinding to MG, but also displayed 83.0% and 87.5% of cross-reactivity with LMG and CV, demonstrating that MIPs could be used as group recognition sorbents in solid phase extraction. The extraction conditions of MISPE column for MG were optimized. Tap water samples spiked with MG at concentration of 0.5-10 ng mL⁻¹ were extracted by MISPE column and analyzed by high performance liquid chromatography. The recoveries of MISPE column for MG extraction were found to be 76.8-93.7% with the relative standard deviations of 2.12-10.09%, indicating the feasibility of the prepared MIPs for MG extraction. No detectable MG was observed in one fish farming water sample and two fish feed samples; while the MG concentrations in two pet fishpond water samples were found at 1.50 ng mL⁻¹ and 0.67 ng mL⁻¹, respectively.     

Characterisation and application of molecularly imprinted polymers for group-selective recognition of antibiotics in food samples

Group-selective molecularly imprinted polymers (MIPs) for amphenicol antibiotics, including chloramphenicol (CAP), thiamphenicol (TAP), florfenicol (FF), and florfenicol amine (FFA), were developed for the first time using TAP as the template molecule. The characteristics of the obtained MIPs were systematically evaluated by chromatographic methods and frontal analysis, demonstrating that the MIPs had excellent chromatographic behaviors, good selectivity, and high-binding capability. A molecularly imprinted solid-phase extraction (MISPE) procedure was developed based on the chromatography results. The MIPs exhibited better group selectivity for CAP, TAP, FF, and FFA than non-imprinted polymers (NIPs) under the optimized washing conditions of 10% acetonitrile in PBS buffer (25 mmol L(-1), pH = 5). Compared with conventional solid-phase extraction, significant recoveries ranging from 92.4% to 98.8% with lower relative standard deviation values in the range of 3.2-7.3% for both intraday- and interday-assays were obtained. The limits of detection (LODs) of MISPE for CAP, TAP, FF, and FFA in shrimp were found to be 0.016, 0.093, 0.102, and 0.029 μg kg(-1), respectively. The results acquired in this study contribute to the strategic development of MIPs and MISPE methods for the multi-residual recognition of antibiotics from complex matrices.     

Citrinin selective molecularly imprinted polymers for SPE

Molecularly imprinted polymers (MIPs) for citrinin (Cit) with 1‐hydroxy‐2‐naphthoic acid (HNA) as mimic template were prepared and the molecularly imprinted SPE method was developed for the detection of Cit in rice with HPLC. The adsorption properties of HNA and Cit on the MIPs and nonimprinted polymers were investigated. It proved that MIPs showed higher selectivity adsorption to HNA and Cit than nonimprinted polymers were. The recoveries of Cit in rice were in the range of 86.7–97.7%. The spiked rice samples and five rice samples in Beijing market were detected using molecularly imprinted SPE method and satisfied results were obtained as discussed in this article.     

Molecularly imprinted polymers for 5-fluorouracil release in biological fluids

The aim of this work was to investigate the possibility of employing Molecularly Imprinted Polymers (MIPs) as a controlled release device for 5-fluorouracil (5-FU) in biological fluids, especially gastrointestinal ones, compared to Non Imprinted Polymers (NIPs). MIPs were synthesized using methacrylic acid (MAA) as functional monomer and ethylene glycol dimethacrylate (EGDMA) as crosslinking agent. The capacity of the polymer to recognize and to bind the template selectively in both organic and aqueous media was evaluated. An in vitro release study was performed both in gastrointestinal and in plasma simulating fluids. The imprinted polymers bound much more 5-Fu than the corresponding non-imprinted ones and showed a controlled/sustained drug release, with MIPs release rate being indeed much more sustained than that obtained from NIPs. These polymers represent a potential valid system for drug delivery and this study indicates that the selective binding characteristic of molecularly imprinted polymers is promising for the preparation of novel controlled release drug dosage form.     

Evidence for shape selectivity in non-covalently imprinted polymers

The existence of shape selectivity in non-covalent molecularly imprinted polymers (MIPs) has been proven using molecular probes. Twelve secondary amines with different sized side chains were imprinted, and enantioselectivity evaluated by HPLC for each amine on each imprinted polymer. Trends in the quantitative structure-binding relationships (QSBR) revealed two major contributions of cavity structure on selectivity afforded by molecularly imprinted polymers. First, sterics play a dominant role in cases where a molecules structure is too big too fit into an imprinted site formed from a smaller template molecule; e.g. on MIPs made with small templates, large analytes give separation factors (α) close to 1.0 (no selectivity), while small analytes give α values of 1.4. Second, molecular structures that are equal to or smaller than those of the template molecule are selected by maximizing Van der Waals interactions within the MIP binding site. Thus, MIPs made with large analytes give α values up to 2.5, while small analytes on the same MIPs give α values closer to 1.1. Template structure also has an effect on MIP enantioselectivity; e.g. branched structures exhibit a 1.7-fold improvement in separation factors (α) by MIPs made for isopropyl versus propyl derivatives, and cyclohexyl versus hexyl derivatives. Full details of these trends are provided in the text.     

分子印迹聚合物固相萃取研究进展

对最新报道的分子印迹聚合物作为固相萃取剂及其在色谱样品前处理方面的应用进行综述和展望,主要包括固相萃取、基质固相分散萃取、固相微萃取、搅拌棒吸附萃取和磁性材料萃取,同时总结了分子印迹聚合物制备技术面临的挑战和问题,提出了可能的解决方案。