Polymer imprinting

Each racemate was applied on a polymer (ca. 0.1. imol per gram dry polymer) imprinted with one antipode of the racemate. Tlie standard mobile phase, consisting of acetonitrile containing various amounts of acetic acid, was used in most cases. Cbz = Carbobenzyloxy, Boc = t-butyloxycarbonyl. 

PMAA refers to polymers imprinted with respective base using MAA as functional monomer. 

Molecular Polymer Imprints (University of Texas) -DNA-Based Identification... 

Hua, E et al. 2004. Polymer imprint lithography with molecular-scale resolution. Nano Lett. 4 2467-2471. 

Not only chiral separations have been achieved with Mi-stationary phases. It has also been demonstrated that the MIP could distinguish between ortho- and para-isomers of carbohydrate derivatives. For example, a polymer imprinted with o-aminophenyl tetraacetyl P-D-galactoside was used to analyze a mixture of p-and o-aminophenyl tetraacetyl P-D-galactoside. As expected, the imprinted ortho analyte eluted after the non-imprinted para component see Fig. 5. Although baseline separation was not obtained, a separation factor of a = 1.51 was observed [19]. 

An interesting example of MIP-LC analytics was presented in a paper, which focused on the separation of antibiotics of similar structures. Columns are (commercially) available to separate penicillins ( 3-lactams) from other antibiotics. However, if the quantification of each of the 3-lactam compounds is required, a more selective stationary phase has to be found. Molecular imprinting allows the fabrication of phases specifically for each 3-lactam. If for instance the concentration of the P-lactam oxacillin in a food sample has to be selectively determined, a polymer imprinted with oxacillin is the right choice. Compared to a standard stationary phase, which only allowed the separation of the entire group of (5-lactams from other non-(3-lactam analytes (e. g., bacitracin), the MIP enables the separation of the imprinted species from the pair of non-imprinted 3-lactams penicillin V and penicillin G see Fig. 6 [29,30]. 

Fig. 5. Separation of (1) para- and (2) ortho-aminophenyl tetraacetyl 3-D-galactoside, using a polymer imprinted with o-aminophenyl tetraacetyl p-galactoside. Reprinted with permission from Nilsson KGI, Sakguchi K, Gemeiner P, Mosbach, K (1995) J Chromatogr 707 199. Copyright 1995 Elsevier Science...

Beside the use of MIPs in conventional HPLC, Mi-polymers may also be established in supercritical fluid chromatography, which is characterized by faster equilibration times combined with the use of the environmental friendly C02 as mobile phase. Although preliminary results show relatively broad peaks, chiral separation could be performed based on polymers imprinted with an enantiomer. However, the long-term stability of the photochemically generated polymers seems to be a problem [89]. 

In a different approach, Lin et al. have used particles derived from a ground MI-bulk polymer and mixed with a polyacryl amide gel for chiral separation. Using a polymer imprinted with L-phenylalanine, D-phenylalanine could be separated from the template with a separation factor of 1.45 [35]. Although the combination of MIP with capillary electrochromatography is still not widely used, the ability to separate enantiomers in nanoliter samples promises interesting developments for the future. 

Fig. 9a, b. Chromatograms obtained after pre-concentration of a 100 ml groundwater sample spiked at 1 p.g 1 1 through a a CP-cartridge b a cartridge filled with a polymer imprinted with terbuthylazine. Peaks 1 = deisopropylatrazine, 2 = deethylatrazine, 3 = simazine, 4 = atrazine, 5 = propazine, 6 = terbuthylazine, I.S. = internal standard (diuron). Reprinted with permission from Ferrer I, Lanza F, Tolokan A, Horvath V, Sellergren B, Horvai G, Barcelo D (2000) Anal Chem 72 3934. Copyright 2000 American Chemical Society... 

Quite a number of recent papers have concentrated on techniques which allow the gravimetric detection of only a few femtograms of an analyte [105]. In one of these, PAHs could be measured in concentration of down to a few ng ml-1 in degraded oil when a piezoelectric quartz crystal coated with Mi-polymers imprinted either with a fresh or with a degraded lubricant was immersed into the... 

Fig. 15. Displacement of radio-labeled analyte analog binding to MI spheres under equilibrium condition. B/B0 is the ratio of the amount of radio-labeled ligand bound in the presence of the displacing ligand (analyte), B0. Displacement of [8-3H]theophylline binding to a polymer imprinted with theophylline. Displacing ligands theophylline ( ) theobromine (+) xanthine (o) caffeine (A). Reprinted with permission from Ye L, Cormack PAG, Mosbach K (1999) Anal Commun 36 35. Copyright 1999 The Royal Society of Chemistry...

Comparable results were presented when the number of Mi-polymers was extended to other templates of similar steroid libraries. The use of a polymer imprinted with 11-deoxycortisol in a separation of 11-a-hydroxyprogesterone, progesterone and 11-deoxycortisol resulted in a chromatogram where the 11-de-oxycortisol eluted last (Fig. 17, bottom). When an 11-a-hydroxyprogesterone imprinted polymer was used, on the other hand, the 11 -a-hydroxyprogesterone was found to elute last (Fig. 17, top) [80]. 

Figure 15.11 Sensor response to herbicide concentration obtained with a polymer imprinted with atrazine. Reprinted from Sergeyeva et al. (1999). Copyright 1999 Elsevier Science.

Separation of theophylline and caffeine by column containing polymer imprinted with theophylline. Caffeine washed right through with CHCI3 solvent. Addition of 20 id. of CH3OH to the solvent breaks hydrogen bonds between theophylline and the polymer and elutes theophylline in a volume of t mL [From W. M. Mulleti and P C. Lai. Anal. Chem. 1998, 70.3636.]... 

Liquid fluorocarbon was used as continuous phase by Perez-Moral and Mayes [19] as well. They proposed a new method for rapid synthesis of MIP beads, in that they prepared 36 polymers imprinted for propranolol and morphine with different amounts of EDMA as a cross-linker and different functional monomers (MAA, acrylic acid, hydroxyethyl methacrylate, 4-vinylpyridine) directly in SPE cartridges. The properties of MIP microspheres prepared by this method were very similar in terms of size, morphology and extent of rebinding to microspheres prepared by conventional suspension polymerisation in perfluorocarbons as well as to bulk polymers prepared in the same solvent. The most notable advantages of this method are no waste production (no transfer of beads during washing steps) and possible direct use for a variety of screening, evaluation and optimisation experiments. 

The Mosbach group described in 1993 the first assay in which polymers imprinted with either theophylline or diazepam replaced antibodies in competitive RILA... 

Surugiu et al. [67] have introduced an Enzyme Immuno-Like Assays (EzILA) for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The label was a 2,4-D conjugate with the tobacco peroxidase (TOP) enzyme, which allows for both colorimetric and chemiluminescent detection. In this case, the polymer imprinted with 2,4-D was synthesized in the form of microspheres. In contrast, despite its higher binding capacity for radiolabeled 2,4-D, a conventional MIP prepared by bulk polymerization showed only weak binding of the 2,4-D-TOP tracer. 

Scheme 2 Polymer imprinted with the structure (6) were able to convert the achiral substrate (7) in the chiral substrate (8)...

The first reported attempt of using MIPs to control the stereochemical course of a reaction dates back to 1980, when the two research groups of Neckers and Shea published, simultaneously, examples of bulk polymers able to control the formation of the product by using a chiral template. Shea et al. reported that bulk polymers imprinted with stereochemically pure ( )-/ra/w-l,2,cyclobutane-dicarboxyilic acid (6) were able to keep a molecular memory of the asymmetry of the template [8]. In fact, this was transferred to an achiral substrate, such as fumaric acid (7), inducing a diastereoselective methylation, which led to trans-1,2,cyclopropane-dicarboxyilic... 

Neckers et al. reported the synthesis of bulk polymers imprinted with one of the three possible isomeric products of the photodimerisation of /ram-cinnamic acid... 

Mosbach and collaborators, in 2001, described an innovative approach to drug discovery using polymers imprinted with a biologically active template [31]. The approach, called anti-idiotypic for the similarity with anti-idiotypic antibodies in the immune-response, can be used to create synthetic receptors able to generate inhibitors or receptor antagonists by exploiting the complementarity with the cavity. The imprinted cavity promotes preferentially the formation of compounds with high affinity, which can later be evaluated for the inhibitory activity and the more active selected for further analysis. 

The authors reported the preparation of polymers imprinted with kallikrein (47) as a template, a known tissue inhibitor, which, after template removal, were used to allow direct coupling between a di-chloro-triazine (48) and a series of aromatic amines. The first substrate was used to resynthesise the compound that was used as template and the yield of this was four times higher than with the corresponding control polymer. Moreover, the same reaction performed in free solution in the same conditions gave no product. The polymer was then tested against the other substrates to verify whether it was possible to synthesise compounds with slightly different properties. This experiment resulted in compounds with lower yields, 21 % for one and 34% for another, whilst using a bulkier substrate it was not possible to obtain any product. 

One of the most investigated type of reaction in the field of catalytic imprinted polymers, as indicated by the large number of publications available, is certainly ester hydrolysis. In particular, a great deal of work has been carried out on systems inspired by hydrolytic enzymes since 1987. In 2000, Shea et al. [37] reported the preparation of enantioselective imprinted polymers for the hydrolysis of N-tert-butoxycarbonyl phenylalanine-p-nitrophenyl ester (55), using a system already developed by the same group in 1994 [19]. The system was inspired by the natural hydrolytic enzyme chymotrypsin and polymerisable imidazole units (27) were used as functional monomers coupled via ester linkages to a chiral phosphonate (56), analogue of (d)- or (L)-phenyl-alanine. After template removal, the imprinted polymers showed selectivity towards the hydrolysis of the enantiomer with which they were imprinted. The ratio of the rate constants, k /k, was 1.9 for the polymer imprinted with the D-enantiomer and kjku was 1.2 for that imprinted with the L-enantiomer. Moreover, the imprinted polymer showed a 2.5-fold increase in the rate of the reaction when compared with the control polymer, imprinted with a... 

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