Abstract
This research project has investigated the feasibility of using immunotechniques for the immuno-detection and monitoring of pesticides in different matrices, mainly water residues. The maximum admissible concentration (MAC) for a single pesticide (0.1μg/L), which was set by the European community, has been successfully detected by the use of immunoassay methods. Competitive enzyme-linked immunosorbent assays (ELISA) suitable for the determination of the urea herbicides isoproturon, chlortoluron and 2,4-D in different types of water and biological fluids have been developed. The production of highly specific antibodies utilised in this work was achieved by covalently coupling thyroglobulin protein with the compound or a synthesised hapten. As a result, the cross-reactivities of the antisera with a range of related and unrelated pesticides was shown to be negligible. Furthermore, the limits of detection of the isoproturon and chlortoluron ELISA methods were 0.03 and 0.015 μg/L respectively, well below the MAC for pesticides in drinking water, whilst the ELISA assay limit of detection for 2,4-D was 50 μg/L which would be sensitive enough for monitoring 2,4-D in air for occupational exposure studies. In addition, reproducible and quantitative recoveries of isoproturon, chlortoluron and 2,4-D from water, obtained from various sources, and biological fluids were possible without any sample preparation requirements. The feasibility of using a luminescent assay with a photographic end point has shown considerable potential by providing a rapid, simple and portable means of monitoring multiple water samples for the presence of pesticides. The assay was able to identify samples containing a pesticide or a class group of pesticides at or above the MAC level. Consequently, this would greatly facilitate the increased monitoring of water supplies in other areas of environmental analysis. A further study was carried out to demonstrate that screening pesticides qualitatively with non-instrumental immunoaffinity columns based on the principles of affinity chromatography and enzyme immunoassay was possible. The method produced qualitative results in 20 min by determining samples containing certain concentrations of pesticides and above as positive samples (i. e. 0.12 μg/L or above for isoproturon and 0.11 μg/L or above for chlortoluron). Samples with a concentration of pesticides below these levels would be considered as negative (no pesticides present at these limits of detection). The advantages of this procedure are that it requires no sample dilutions, separation steps, or precise timing (except for reading colour). Moreover, the technique characteristics made it ideally suited for monitoring pesticides on-site in a different range of matrices which could be performed by non-skilled personnel thereby reducing the workload involved in meeting EC regulations. The data reported in this thesis demonstrate the potential use of immunoassays in environmental analysis potentially offering a substantial improvement to the monitoring programmes of pesticides by providing tests which are sensitive, rapid, inexpensive and simple to perform. This should encourage further work on the development of immunoassays for other pesticides and for toxic or hazardous chemicals in general.