Determination of non-extractable triazine residues by enzyme immunoassay: Investigation of model compounds and soil fulvic and humic acids

2-Chloro-4-arylamino-6-alkyl-1,3,5-triazines were used as model compounds for non-extractable (bound) triazine residues. They were tested as targets in a competitive enzyme immunoassay with polyclonal and monoclonal antibodies originally raised against atrazine. Cross-reactivities were determined as a measure for the affinity of antibodies toward their respective analytes. 2-Chloro-8-isopropyl derivatives were recognized as well as free atrazine (70-148% cross-reactivity, atrazine = 100%) with the exception of 2-chloro-4-[2'-carboxy-4',5'-dimethoxyanilino]-6- isopropylamino-s-triazine, carrying a carboxyl group in the arylamino side chain (<0.1-9% cross-reactivity). Functional group variation of positions 2 and 8 of the triazine ring resulted in cross-reactivities less than 15%. Arylamino-s-triazine samples from photolytic degradation experiments were analyzed by immunoassay and HPLC. Similar concentrations were obtained by both methods. Then the enzyme immunoassay was applied for the screening of triazines bound to natural fulvic and humic acids. Antibodies with different binding properties were used. It was possible to determine non-extractable atrazine residues in fulvic and humic acids isolated from agricultural soils. Up to 50 ng of bound residues per g of soil was found. Based on the binding by different antibodies, a proposal for the ligation of non-extractable atrazine to soil particles is made.