Imprinted site

In the elucidation of retention mechanisms, an advantage of using enantiomers as templates is that nonspecific binding, which affects both enantiomers equally, cancels out. Therefore the separation factor (a) uniquely reflects the contribution to binding from the enantioselectively imprinted sites. As an additional comparison the retention on the imprinted phase is compared with the retention on a nonimprinted reference phase. The efficiency of the separations is routinely characterized by estimating a number of theoretical plates (N), a resolution factor (R ) and a peak asymmetry factor (A ) [19]. These quantities are affected by the quality of the packing and mass transfer limitations, as well as of the amount and distribution of the binding sites. 

Eventually, a matrix is formed containing the molecularly imprinted sites locked into a matrix such as illustrated in the following structure ... 

Fig. 1 General principle of molecular imprinting. A molecular template (T) is mixed with functional monomers (M) and a cross-linker (CL) resulting in the formation of a self-assembled complex (1). The polymerization of the resulting system produces a rigid structure bearing imprinted sites (2). Finally removal of the template liberates cavities that can specifically recognize and bind the target molecule (3). Adapted with permission from [3]. Copyright 2003 American Chemical Society...

More recently, Priego-Capote et al. reported on the production of MIP nanoparticles with monoclonal behaviour by miniemulsion polymerisation [63]. In the synthetic method that they employed, they devised to use a polymerisable surfactant that was also able to act as a functional monomer by interacting with the template (Fig. 4). The crosslinker content was optimised at 81% mol/mol (higher or lower contents leading to unstable emulsions). In this way, the authors were able not only to produce rather small particles (80-120 nm in the dry state) but also to locate the imprinted sites on the outer particle surface. The resulting MIP nanobeads were very effective as pseudostationary phases in the analysis of (/ ,S)-propranolol by CEC. 

Fig. 10 Schematic description of the strategy followed by Whitcombe et al. for the preparation of core-shell MIP nanoparticles with imprinted sites located on the particle surface. Reproduced with permission from [124]...

In an alternative approach, MIP membranes can be obtained by generating molec-ularly imprinted sites in a non-specific matrix of a synthetic or natural polymer material during polymer solidification. The recognition cavities are formed by the fixation of a polymer conformation adopted upon interaction with the template molecule. Phase inversion methods have used either the evaporation of polymer solvent (dry phase separation) or the precipitation of the pre-synthesised polymer (wet phase inversion process). The major difficulties of this method lay both in the appropriate process conditions allowing the formation of porous materials and recognition sites and in the stability of these sites after template removal due to the lack of chemical cross-linking. 

The study of QDs and MIPs in the development of the detection scheme for pyrethroids has very recently been reported [72]. Pyrethroids are pesticides commonly used against insects being widely used in agriculture, medicine, industry and the household. The QDs were embedded in silica nanospheres having lambda-cahalothrin imprinted sites on their surfaces. As also presented in the previous example, the principle of recognition was based on a decrease of fluorescence with an increase of analyte concentration. The lambda-cahalothrin imprinted... 

Moreover, the higher the concentration of the imprinted sites in the MIP film the lower, advantageously, is LOD. For instance, LOD for the MIP films fabricated from acrylic polymers is, unfortunately, very high. This is because their sensing capabilities are hindered due to a relatively low concentration of recognition sites. Therefore, other different routes of MIP fabrication, using other than acrylic monomers, have been developed to increase this concentration. 

Whitcombe et al. observed a strong decrease of the cholesterol concentration, and hence excellent binding for cholesterol with p(EGDMA-co-CVPC) shell/ pMMA core particles in isohexane. When the carbonate ester was not hydrolyzed and hence no imprinted sites existed in the shell, only 2.7 pmol cholesterol per g particles were bound corresponding to the non-specifically adsorbed templates. Slightly less cholesterol was adsorbed to particles prepared without CVPC at all thus formed with a pure pEGDMA shell (<2 pmol g-1). 

The matrix polymer has to be rather rigid to conserve the empty imprinted sites after extraction of the template, this is achieved by extensive crosslinking. Generally, the crosslinker content of a mixture has to exceed a threshold value (>40%) to obtain selectivity [444,447]. Further, the chemical structure of the polymer network is of minor importance for the selectivity, though its polarity should be adapted to the imprinting molecule [444]. To obtain a maximum number of accessible sites,... 

It is of obvious importance that the functional monomers strongly interact with the template prior to polymerisation, since the solution structure of the resulting assemblies presumably defines the subsequently formed binding sites. By stabilising the monomer-template assemblies it is possible to increase the number of imprinted sites. At the same time the number of non-specific binding sites will be minimised, since there will be a reduction in the amount of free, non-associated functional monomer. For any particular template, the following factors that are likely to affect the recognition properties of the site have been identified (Fig. 5.14). 

As noted in the first proposed explanation (a) for the change in enantioselectiv-ity, an increase in pH may per se lead to a decrease in the enantioselectivity of the imprinted sites. One way to separate these effects is to measure the complete isotherms at several pH values and evaluate how the relative population of nonspecific and specific sites, as well as their affinity for the template, change with respect to the mobile phase pH [40,50]. 

---