Determination of Non-extractable Triazine Residues by Enzyme Immunoassay:  Investigation of Model Compounds and Soil Fulvic and Humic Acids

Abstract

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-6-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 6 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.

Approaches to the Immunochemical Analysis of Non-extractable Triazine Residues in Refractory Organic Substances (ROS) and Characterization of ROS

“…2) [16,28]. The same Ab binding pattern was observed when FA and HA from soil containing non-extractable residues were investigated by EIA [17].…”

“…Sensitivities are reached down to 10...20 ng/L [15,16], and only small sample volumes (a few milliliters) are necessary. Recently, EIA also have been applied for the determination of non-extractable atrazine residues in soil [17,18].…”

“…The methanolic eluate was diluted 1:100 with bidistilled water prior to the test. The assays were performed as described by Dankwardt et al [17]. b Total amount of triazines bound to the ROS in one dialysis tubing (e.g., 30 ng atrazine bound to 3 mg DOC: experiment atrazine, pH 9), corrected for the dilution factor by dialysis, devided by total amount of free triazines from the incubation solution (20 µg in 200 mL (100 µg/L)) × 100, in %.…”

Atrazine interaction with tropical humic substances by Enzyme Linked Immunosorbent Assay

“…Some pesticides can be adsorbed or covalently bound to the humic polymer leading to erroneous results. On the other hand, humic substances if present at high concentrations may exhibit nonspecific binding properties in the ELISA [22]. In this study we have used the ELISA developed in our laboratory that has shown a good rep ro d u c i b i l i t y, and the coefficient of variation the IC 50 (concentration reducing the signal to 50% of the control) was less than 10%.…”

Spectroscopic Characterization of Fulvic Acid Fractions of a Contaminated Groundwater

“…The main part in NOM can be attributed to humic substances (HS) which are divided into fulvic acids (FA) and humic acids (HA) on an operationally-defined basis [2,3]. Different mechanisms for the interaction between NOM and organic contaminants ranging from weak van der Waals forces to covalent binding of the contaminant to the NOM matrix were reported [4][5][6]. Fluorescence spectroscopy is a multidimensional technique with three main independent variables: excitation wavelength λ ex , emission wavelength λ em , and fluorescence decay time τ f .…”