Catalytically active polymers obtained by molecular imprinting and their application in chemical reaction engineering.

Molecular imprinting is a way of creating polymers bearing artificial receptors. It allows the fabrication of highly selective plastics by polymerizing monomers in the presence of a template. This technique primarily had been developed for the generation of biomimetic materials to be used in chromatographic separation, in extraction approaches and in sensors and assays. Beyond these applications, in the past few years molecular imprinting has become a tool for producing new kinds of catalysts. For catalytic applications, the template must be chosen, so that it is structurally comparable with the transition state (a transition state analogue, TSA) of a reaction, or with the product or substrate. The advantage of using these polymeric catalysts is obvious: the backbone withstands more aggressive conditions than a bio material could ever survive. Results are presented showing the applicability of a molecularly imprinted catalyst in different kinds of chemical reactors. It is demonstrated that the catalysts can be utilized not only in batch but also in continuously driven reactors and that their performance can be improved by means of chemical reaction engineering.

Food analyses using molecularly imprinted polymers.

Molecular imprinting technology (MIT) is a technique for generating polymers bearing biomimetic receptors. It offers several advantages to the agrofood industry in areas such as analysis, sensoring, extraction, or preconcentration of components. It has the potential of becoming a tool for acquiring truly simple, rapid, and robust direct measurements. In this review, the special features of MIT that have bearing on food science and technology are highlighted.

New configurations and applications of molecularly imprinted polymers.

Molecularly imprinted polymers (MIPs) are applicable in a variety of different configurations. For example, bulk polymers imprinted with beta-lactam antibiotics are presented to be used as stationary phases for the chromatographic separation of beta-lactam antibiotics with both aqueous and organic mobile phases. However, in some analytical applications, monosized spherical beads are preferred over the currently used ground bulk polymers. A precipitation polymerization technique allows preparation of monosized spherical imprinted beads with diameters down to 200 nm having excellent recognition properties for different target molecules. Nevertheless, with current imprinting protocols a substantial amount of template has to be used to prepare the polymer. This can be problematic if the template is poorly soluble, expensive or difficult to obtain. It is shown that for analytical applications, the functional monomer:template ratio can be drastically increased without jeopardizing the polymer's recognition properties. Furthermore, a substantial reduction of the degree of crosslinking is demonstrated, resulting in much more flexible polymers that are useful for example the preparation of thin imprinted films and membranes for sensors. Apart from analysis, MIPs also are applicable in chemical or enzymatic synthesis. For example, MIPs using the product of an enzyme reaction as template are utilized for assisting the synthetic reaction by continuously removing the product from the bulk solution by complexation. This results in an equilibrium shift towards product formation.

Analysis of endotoxins by capillary electrophoresis.

Endotoxins are part of the outer membrane of gram-negative bacteria such as E. coli. Upon entering the blood stream, they cause a violent, sometimes life-threatening, response of the immune system. Endotoxins are lipopolysaccharides (LPS), lacking optically active groups, and their detection in the underivatized state can be difficult. In this paper the potential of capillary electrophoresis (CE) for LPS analysis is investigated. By using a standard phosphate buffer method, concentrations down to 100 micrograms/mL can be detected within 6 min. The detection limit can be lowered by one order of magnitude by using a sodium dodecyl sulfate (SDS)/borate buffer, pH 9.2. In this buffer, the SDS serves to homogenize the size of the LPS aggregates, while the borate forms complexes with the diol groups of the molecule, thereby enhancing its optical activity. The formation of LPS-affinity complexes with the UV-active polymyxin B or labeling of the LPS with a fluorophore (fluorescein isothiocyanate) was unsuccessful. Best results, in terms of detection limit and speed, were obtained with an indirect UV-detection CE method. By using a strongly UV-active electrophoresis buffer, endotoxins could be detected as "negative" peaks. In this case, a detection limit of 3 micrograms/mL (35 pM) was determined. Proteins and other UV-active substances did not disturb the assay, since they generated no detectable signals. The indirect UV detection was used to quantify the residual LPS content of a DNA preparation from E. coli.

Analysis of amatoxins alpha-amanitin and beta-amanitin in toadstool extracts and body fluids by capillary zone electrophoresis with photodiode array detection.

Over 90% of the lethal cases of mushroom toxin poisoning in man are caused by a species of amanita. The amatoxins (especially alpha- and beta-amanitin) found in amanita deserve special attention, because of their high pharmacological potency, their high natural concentration and their high chemical and thermal stability. Measures can be taken to improve the survival rates (aggressive gastroenteric decontamination, liver protection therapy) if the poisoning is diagnosed correctly and as early as possible. The standard assay for alpha-amanitin is a radioimmunoassay (RIA). Among other reagents, this assay uses 125I-labelled alpha-amaintin, which has a low shelf life. The assay is therefore not available at all hospitals and all year round. In this paper, a first attempt to employ capillary zone electrophoresis (CZE) to quantify amatoxins alpha- and beta-amanitin in urine samples of afflicted patients and in toadstool extracts is described. Diode array detection is used for identification of the resolved substances in the electropherogram. An analysis requires 20 min. The detection limit is 1 microgram/ml, i.e., 5 pg absolute. Relative standard deviations are between 1 and 2% for the calibration standards (peak height and area) and ca. 7.5% for the real samples. Advantages of the CZE over the RIA include lower cost, the possibility of quantifying several toxins in one analysis, less consumption of potentially harmful reagents (no radio-labelled substances, no addition of alpha-amanitin as reagent) and, most importantly, all-year-round availability of the assay. The detection limit is still somewhat high and does not cover the entire clinically relevant range. Attempts to lower the detection limit by the necessary order of magnitude are currently under way in our laboratory. These include application of laser-induced fluorescence detection, liquid chromatography-CZE and CZE-mass spectrometry techniques.