A molecularly imprinted polymer (MIP) is a polymer that has been processed using the molecular imprinting technique which leaves cavities in the polymer matrix with an affinity for a chosen "template" molecule. The process usually involves initiating the polymerization of monomers in the presence of a template molecule that is extracted afterwards, leaving behind complementary cavities. These polymers have affinity for the original molecule and have been used in applications such as chemical separations, catalysis, or molecular sensors. Published works on the topic date to the 1930s.
Molecular imprinting techniques (state of the art and perspectives) Molecular imprinting is the process of generating an impression within a solid or a gel, the size, shape and charge distribution of which corresponds to a template molecule typically present during polymerisation. The result is a synthetic receptor capable of binding to a target molecule, which fits into the binding site with high affinity and specificity. The interactions between the polymer and the template are similar to those between antibodies and antigens, consisting of electrostatic interactions, hydrogen bonds, Van der Waals forces, and hydrophobic interactions.
One of the greatest advantages of artificial receptors over naturally occurring receptors is freedom of molecular design. Their frameworks are not restricted to proteins, and a variety of skeletons (e.g., carbon chains and fused aromatic rings) can be used. Thus, the stability, flexibility, and other properties are freely modulated according to need. Even functional groups that are not found in nature can be employed in these synthetic compounds. Furthermore, when necessary, the activity in response towards outer stimuli (photo-irradiation, pH change, electric or magnetic field, and others) can be provided by using appropriate functional groups.
In a molecular imprinting processes, one needs a 1) template, 2) functional monomer(s) 3) cross-linker, 4) radical or other polymerization initiator, 5) porogenic solvent and 6) extraction solvent. According to polymerization method and final polymer format one or some of the reagent can be avoided.
Creating polymers There are two main methods for creating these specialized polymers. The first is known as self-assembly, which involves the formation of polymer by combining all elements of the MIP and allowing the molecular interactions to form the cross-linked polymer with the template molecule bound. The second method of formation of MIPs involves covalently linking the imprint molecule to the monomer. After polymerization, the monomer is cleaved from the template molecule. The selectivity is greatly influenced by the kind and amount of cross-linking agent used in the synthesis of the imprinted polymer. The selectivity is also determined by the covalent and non-covalent interactions between the target molecule and monomer functional groups. The careful choice of functional monomer is another important choice to provide complementary interactions with the template and substrates."Characteristic and Synthetic Approach of Molecularly Imprinted Polymer" Int. J. Mol. Sci. 2006, 7, 155–178 In an imprinted polymer, the cross-linker fulfills three major functions: First of all, the cross-linker is important in controlling the morphology of the polymer matrix, whether it is gel-type, macroporous or a microgel powder. Secondly, it serves to stabilize the imprinted binding site. Finally, it imparts mechanical stability to the polymer matrix. From a polymerization point of view, high cross-link ratios are generally preferred in order to access permanently porous materials and in order to be able to generate materials with adequate mechanical stability. The self-assembly method has advantages in the fact that it forms a more natural binding site, and also offers additional flexibility in the types of monomers that can be polymerized. The covalent method has its advantages in generally offering a high yield of homogeneous binding sites, but first requires the synthesis of a derivatized imprint molecule and may not imitate the "natural" conditions that could be present elsewhere. Over the recent years, interest in the technique of molecular imprinting has increased rapidly, both in the academic community and in the industry. Consequently, significant progress has been made in developing polymerization methods that produce adequate MIP formats with rather good binding properties expecting an enhancement in the performance or in order to suit the desirable final application, such as beads, films or nanoparticles.
Challenges One of the key issues that have limited the performance of MIPs in practical applications so far is the lack of simple and robust methods to synthesize MIPs in the optimum formats required by the application. Chronologically, the first polymerization method encountered for MIP was based on "bulk" or solution polymerization. This method is the most common technique used by groups working on imprinting especially due to its simplicity and versatility. It is used exclusively with organic solvents mainly with low dielectric constant and consists basically of mixing all the components (template, monomer, solvent and initiator) and subsequently polymerizing them. The resultant polymeric block is then pulverized, freed from the template, crushed and sieved to obtain particles of irregular shape and size between 20 and 50 μm.
Depending on the target (template) type and the final application of the MIP, MIPs are appeared in different formats such as nano/micro spherical particles, nanowires and thin film or membranes. They are produced with different polymerization techniques like bulk, precipitation, emulsion, suspension, dispersion, gelation, and multi-step swelling polymerization. Most of investigators in the field of MIP are making MIP with heuristic techniques such as hierarchical imprinting method. The technique for the first time was used for making MIP by Sellergren et al. for imprinting small target molecules. With the same concept, Nematollahzadeh et al. developed a general technique, so-called polymerization packed bed, to obtain hierarchically-structured, high capacity protein imprinted porous polymer beads by using silica porous particles for protein recognition and capture.
Solid-phase synthesis Solid-phase molecular imprinting has been recently developed as an alternative to traditional bulk imprinting, generating water-soluble nanoparticles. As the name implies, this technique requires the immobilisation of the target molecule on a solid support prior to performing polymerisation. This is analogous to solid-phase synthesis of peptides. The solid phase doubles as an affinity separation matrix, allowing the removal of low-affinity MIPs and overcoming many of the previously described limitations of MIPs: Separation of MIPs from the immobilised template molecule is greatly simplified. Binding sites are more uniform, and template molecules cannot become trapped within the polymer matrix. MIPs can be functionalised post-synthesis (whilst attached to the solid phase) without significantly influencing binding sites. The immobilised template can be reused, reducing the cost of MIP synthesis.
MIP nanoparticles synthesised via this approach have found applications in various diagnostic assay and sensors.
An adaptation of the solid-phase protocol was performed by Sullivan et al. who used a modified aptamer as a recognition macromonomer, encapsulated within a polymer nanoparticle scaffold. Producing the first truly aptamer-MIP hybrid (aptaMIP), improving target recognition.
Molecular modelling Molecular modelling has become a convenient choice in MIP design and analysis, allowing rapid selection of monomers and optimization of polymer composition, with a range of different techniques being applied. This application of molecular modelling utilises a method of automated screening of a large database of monomers against a given target or template with a molecular mechanics approach. In recent years technological advances have permitted more efficient analysis of monomer-template interactions by quantum mechanical molecular modelling, providing more precise calculations of binding energies. Molecular dynamics has also been applied for more detailed analysis of systems before polymerisation, and of the resulting polymer, which by including more system components (initiator, cross-linkers, solvents) provides greater accuracy in predicting successful MIP synthesis than monomer-template interactions alone. Molecular modelling, particular molecular dynamics and the less common coarse-grained techniques, can often also be integrated into greater theoretical models permitting thermodynamic analysis and kinetic data for mesoscopic analysis of imprinted polymer bulk monoliths and MIP nanoparticles.
Applications Niche areas for application of MIPs are in sensors and separation. Despite the current good health of molecular imprinting in general, one difficulty which appears to remain to this day is the commercialization of molecularly imprinted polymers. Despite this, many patents (1035 patents, up to October 2018, according to the Scifinder data base) on molecular imprinting were held by different groups.
Fast and cost-effective molecularly imprinted polymer technique has applications in many fields of chemistry, biology and engineering, particularly as an affinity material for sensors, detection of chemical, antimicrobial, and dye, residues in food, adsorbents for solid phase extraction, binding assays, artificial antibodies, chromatographic stationary phase, catalysis, drug development and screening, and byproduct removal in chemical reaction. Molecular imprinted polymers pose this range of capabilities in extraction through specific micro-cavity binding sites. Due to the specific binding site created in a MIP, this technique is showing promise in analytical chemistry as a method for solid phase extraction. The capability for MIPs to be a cheaper easier production of antibody/enzyme-like binding sites doubles the use of this technique as a breakthrough in medical research and application. Such possible medical applications include controlled release drugs, drug monitoring devices, and biological receptor mimetics. Additionally, MIPs can act as enzyme inhibitors by selectively binding to an enzyme's active site or key allosteric region, blocking substrate access and effectively reducing the enzyme's catalytic activity. Beyond this, MIPs show a promising future in the developing knowledge and application in food sciences.Sensor Laboratory CNR-IDASC & University of Brescia. Biosensors. (accessed Feb, 29 2012)
The binding activity of MIPs can be lower compared than that of specific antibodies, even though examples have been reported of MIPs with comparable or better performance to commercially produced antibodies. This yields a wide variety of applications for MIPs from efficient extraction to pharmaceutical/medical uses. MIPs offer many advantages over protein binding sites. Proteins are difficult and expensive to purify, denature (pH, heat, proteolysis), and are difficult to immobilize for reuse. Synthetic polymers are cheap, easy to synthesize, and allow for elaborate, synthetic side chains to be incorporated. Unique side chains allow for higher affinity, selectivity, and specificity.
Molecularly imprinted assays