Development of Immunoenzyme Assay of Herbicide Acetochlor and Its Application to Soil Testing with Comparison of Sample Preparation Techniques

Acetochlor is a selective herbicide affecting weeds of cereal plants. Its analysis in soils allows accessing their suitability for crops and risks of contamination of agricultural products. The aim of this study was to develop a microplate enzyme immunoassay for the determination of acetochlor in soil extracts. For the development, rabbit antibodies specific to acetochlor were obtained by immunization with a conjugate of carrier protein with a derivative of acetochlor with mercaptopropionic acid. Another derivative with mercaptosuccinic acid was applied for immobilization on the solid phase. In the study, organic extracts have been obtained from soil varying solvents and their ratios, and using QuEChERS protocol. The extracts have been tested to estimate residual influences of the sample matrix. Optimal conditions for the immunoassay were selected, appropriate sample preparation techniques, and the composition of the medium for competitive immune interaction. The most effective approach involved dichloromethane extraction, followed by careful evaporation and subsequent reconstitution of the dry residue in a 10 mM phosphate-buffer solution supplemented with 0.1% gelatin. The resulting analytical system exhibited a detection limit of 59.4 ng/mL for acetochlor, with a working range spanning from 112 to 965 ng/mL. Taking into account the soil sample preparation, the LOD was estimated as 0.3 µg/g with the working range from 0.66 to 5.7 µg/g of soil. Analysis of prepared extracts from gray forest soil demonstrated a revealing of acetochlor between 74% and 124%.