Print molecule

By analogy, a great many of other functionalized styrenes, including carboxyHc acids, amino acids, Schiff bases, or specific compounds, eg, l-DOPA, have successfully been appHed as print templates. Moreover, it has also been shown that siUca gel can be imprinted with similar templates, and that the resulting gel has specific recognition sites determined by the print molecule (162—164). 

Fig. 6. A Chromatogram of a mixture containing the print molecule (oxacillin), two other p-lactam-antibiotics (penicillin G and penicillin V) and a non- 3-lactam-antibiotic (bacitracin) on an oxacillin imprinted MIP containing 4-vinylpyridine residues, cross-linked with TRIM. The analysis was performed in organic mobile phase (ACN/AcOH,99 l).B Same conditions but using the respective non-imprinted control polymer. C Structures of penicillin V, penicillin G, and oxacillin. Reprinted with permission from Skudar K, Briiggemann O, Wittelsberger A, Ramstrom O (1999) Anal Commun 36 327. Copyright 1999 The Royal Society of Chemistry...

If a polymer is prepared in the presence of molecules, the "print molecules" of which are extracted after polymerization, the remaining polymer may contain cav hies, prints, or footprints that can recognize the print molecule. Actually, the cast relates to the matrix molecule like lock-and-key til. (see Fig. 11. 

The print molecule (e.g. clenbuterol) is dissolved in a porogen (e.g. acetonitrile) together with either one or two monomers (e.g. methacrylic acid and 4-vinylpyridine). This allows non-covalent complexes to form... 

The final step is to extract the print molecule before the MIP particles can be used as solid phase for separation and concentration studies. A range of extractants have been used in this step, e.g. methanol containing triethylamine or acetic acid are added until a steady baseline is obtained. 

Fig. 4.5 Chromatogram representing print molecule (oxacillin) and two other /3-lactam antibiotics (penicillin G and V) on oxacillin MIP (a) compared with the control blank MIP (b). Reprinted from J. Chromatogr. A, 889, Briiggemann et al. New configurations and applications of molecularly imprinted polymers , 15-24 (2000), with permission...

MIPs used as chiral stationary phases in o-CEC, p-CEC as well as in rod-CEC have shown high selectivity but relatively low efficiency. Most of the reported enantiomer separations on these phases were performed without pressurization of the flow system. Only Schweitz et al. described on the enantiomer separation of propranolol and metoprolol (print molecule R-propranolol or S-metoprolol) [57] and ropivacaine, mepivacaine and bupivacaine (print molecule S-ropivacaine) [58] by... 

Conventionally, MlPs are obtained by bulk co-polymerization from a mixture consisting of a functional monomer, cross-linker, chiral template, and a porogenic solvent mixture. Nowadays, imprinting via non-covalent template binding is preferred over the covalent mode and involves three major steps (see Fig. 9.9). (i) Functional monomers (e.g. methacrylic acid, MAA) and a cross-linker (e.g. ethyleneglycol dimethacrylate, EDMA) assemble around the enantiomeric print molecule, e.g. (S)-phenylalanine anilide (1), driven by non-covalent intermolecular interactions, e.g. ionic interactions, hydrogen bonding, dipole-dipole interaction. Tr-rt-interaction. (ii) By thermally or photochemi-... 

Although molecular imprinting is a fascinating tool for tailoring the enantioselectivity of a CSP, from a practical standpoint MlP-type CSPs are problematic for analytical applications. This is mainly due to (i) their poor efficiency, in particular for the high-affinity enantiomer and print molecule, and (ii) the limited range of applicability, i.e. only for the racemate of the print molecule and structurally closely related SAs for which cross-selectivity exists. These major limitations are the main reasons why there are no MlP-type CSPs currently available on the market. 

Zn(II) was employed as a print molecule because of its strong interaction with the bifunctional monomer, DDDPA. Divinylbenzene, L-glutamic acid dioleylester ribitol and toluene were used as matrix-forming monomer, emulsion stabiliser and diluent, respectively. After polymerisation, the print molecules were removed from the resin, upon which selective recognition sites were formed. The schematic illustration of surface template polymerisation with DDDPA is shown in Scheme 9.8. The Zn(II)-imprinted resins were ground into particles, whose volume-averaged diameters were ca. 40 pm. The yield was ca. 80%. 

In the covalent approach, the print molecules are covalently coupled to polymerisable molecules prior to polymerisation (see Chapter 4). The covalent bonds have to be relatively easy to break to allow cleavage of the print molecules after polymerisation. Print molecules have been coupled to monomers through the formation of boronic, carboxylic and phosphonic ester bonds, amide bonds, imines and ketals. After copolymerisation with a high degree of cross-linker, the print molecules are cleaved from the polymer. A representative example of the covalent approach is shown in Fig. 17.1. 

Non-covalent molecular imprinting relies on multiple non-covalent interactions between the print molecules and the monomers. The monomers are chosen to allow hydrogen bonds, ionic interactions, n-n interactions and/or hydrophobic interactions with the print molecules. Before polymerisation, the monomers and the print... 

A broad range of functional monomers and cross-linkers has been used for the preparation of MIPs. The choice of the functional monomers depends on the nature and functionalities of the print molecule. The most widely used monomer is methacrylic acid, which has been shown to interact through ionic interactions and hydrogen bonds with amines, amides, carbamates and carboxylic acids [13-15]. The monomers and the print molecules self-assemble upon mixing and the strength of the complex is of importance for the selectivity of the polymer. For this reason, a considerable amount of research effort has focused on finding optimal monomers for various classes of print molecules and functionalities. For example, for some print molecules, polymers prepared with vinylpyridines [16,17], 2-(trifluoromethy-l)acrylic acid [18] or acrylamide [19] resulted in higher selectivities and affinities than polymers made from methacrylic acid. Mixtures of functional monomers... 

Not only the rigidity is crucial to the efficiency of MIPs, but also the accessibility as many recognition sites as possible should be accessible for rebinding. The material should therefore be porous. This is realised by dissolving monomers, cross-linkers and print molecules in a porogenic solvent prior to polymerisation. The effect of the solvent on the polymer morphology can be monitored by measuring physical parameters such as surface area, pore diameter and pore volume. 

Initial studies on non-covalent MIPs, pioneered by Mosbach and co-workers, focused mainly on the preparation of materials selective for amino acid derivatives [13-17,19 23,30-38]. The polymers did not only possess selectivity for the amino acid used as the print molecule, but were also found to be enantioselective the... 

The imprinting effects of MIPs prepared with optically active compounds as the print molecules are readily demonstrated by chromatographic evaluations. For example, when the L-enantiomer of an amino acid derivative is used as the print species, a column packed with the resulting polymer will retain the L-enantiomer longer than the o-enantiomer and vice versa when the o-enantiomer is used as the print molecule. Reference polymers prepared with the racemate or without print molecule will not be able to resolve the racemate. A steroselective memory is hence induced in the polymers by the print molecules and is in many cases very precise. 

The print molecules and their optical antipodes were separated. 

A comparison of six different CSPs, all imprinted with the same print molecule... 

The polymers were also able to resolve racemic naproxen, but not the racemates of ibuprofen and ketoprofen (Fig. 17.9). Even though direct comparisons of the two chromatograms cannot be done because of differing flow rates, it is not obvious that the chromatographic efficiency was better with the uniformly sized beads (Fig. 17.9b) than with the irregular particles (Fig. 17.9a). This may be due to the detrimental influence of water on the stability of the monomer-print molecule complex... 

MIPs selective for phenylalanine anilide have been evaluated as stationary phases in TLC [58], Glass backing plates were coated with mixtures of finely ground MIP particles and binders. The plates showed a preferential retardation of the enantiomer used as the print molecule (Fig. 17.13). The R[ values of both enantiomers on non-imprinted polymers were higher than those observed on the... 

Fig. 17.13. Separation of l- and o-phenylalanine anilide on TLC plates covered with poly(-methacrylic acid-co-EDMA) imprinted with (a) L-phenylalanine anilide, (b) o-phenylalanine anilide and (c) no print molecule. Elution with CH3CN-HOAC (99 5). (Adapted from [58], with permission from the American Chemical Society, USA.)...

Figure 17.14a shows the separation of racemic propranolol by CEC on a capillary column filled with poly(methacrylic acid-co-TRIM), polymerised in situ using (i )-propranolol as the print molecule [61]. Several attractive features make this system look promising for the future very low consumption of the print molecule (in this case only 10 pg), fast preparation of the capillaries (3 h) and fast separation (less than 2 min). In Fig. 17.14b, MIP particles selective for (5)-propranolol were added to the mobile phase in a CE separation [62]. The application of MIPs in CE and CEC is described in more detailed in Chapter 16.

To be able to attribute the binding of an MIP to an imprinting effect it is of utmost importance to show that specific recognition sites have been formed due to the presence of the print molecules during the polymerisation. This is usually done by comparisons with appropriate reference polymers. Polymers prepared without print molecules are not always the best choice, since the physical properties (surface area, porosity, etc.) of these polymers often differ from those of imprinted polymers. Reference polymers prepared with the optical antipode or a racemic mixture as the print species are preferred. The selectivity will be reversed when using the optical antipode and a racemic mixture will give a polymer incapable of separating the two enantiomers (unless the monomer(s) is/are chiral). 

An early attempt to make a real electrochemical sensor based on a molecularly imprinted methacrylate polymer utilised conductometric measurements on a field-effect capacitor [76]. A thin film of phenylalanine anilide-imprinted MAA-EDMA copolymer was deposited on the surface of semiconducting p-type silicon and covered with a perforated platinum electrode. An AC potential was applied between this electrode and an aluminium electrode on the back side of the semiconductor and the capacitance measured as a function of the potential when the device was exposed to the analyte in ethanol. The print molecule could be distinguished from phenylalanine but not from tyrosine anilide and the results were very variable between devices, which was attributed to difficulties in the film production. The mechanism by which analyte bound to the polymer might influence the capacitance is again rather unclear. 

Fig. 21.3. Schematic figure of iton-covalent imprinting of polyurethane with an organic solvent as template and porogen. Highly robust MIPs are best prepared over night at room temperature. Template removal is achieved by evaporation or dissolution. Due to the ultra-thin layers the print molecules are often removed completely.

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