Application of fluorescence polarization immunoassay for acetochlor determination in medicinal plant material

Introduction. Acetochlor is one of the most common herbicides capable of accumulating in medicinal plant raw materials and posing a health risk. A method for acetochlor determination was developed using fluorescence polarization immunoassay (FPIA), a homogeneous and specific technique employed for analysis of low-molecular-weight toxicants.

Aim. To develop and validate a fluorescence polarization immunoassay for quantitative analysis of the herbicide acetochlor in medicinal plant material, exemplified by corn silk (Zea mays L.).

Materials and methods. For the development of the method, polyclonal rabbit antibodies against acetochlor and a tracer consisting of an acetochlor derivative conjugated with ethylenediamine fluorescein (AMPA-EDF) were used. The analysis was performed by fluorescence polarization using a portable Sentry-200 fluorimeter (Ellie LLC, USA). Data processing was carried out with SigmaPlot 11 software package (Systat Software Inc., USA). The study was performed on samples of corn silk medicinal plant material purchased from a local pharmacy and an online store. Method validation included the following parameters: limit of detection, limit of quantification, analytical range, precision, accuracy, and specificity.

Results and discussion. A method for the quantitative determination of the pesticide acetochlor in the herbal medicinal raw material corn silk (Zea mays L.) has been developed, optimized, and validated using fluorescence polarization immunoassay (FPIA). The determination range was from 0.024 to 1.56 mg/kg. The specificity, precision, and accuracy of the developed method were confirmed.

Conclusion. The fluorescence polarization immunoassay (FPIA) method was applied for the quantitative determination of the pesticide acetochlor in herbal medicinal raw material, using corn silk as an example.

1. Lengyel Z., Földényi R. Acetochlor as a soil pollutant. Journal of Soils and Sediments. 2002;2:155. https://doi.org/10.1007/BF02988468

2. Wang W.-G., Li M.-Y., Diao L., Zhang C., Tao L.-M., Zhou W.-X., Xu W.-P., Zhang Y. The health risk of acetochlor metabolite CMEPA is associated with lipid accumulation induced liver injury. Environmental Pollution. 2023;331(2):121857. https://doi.org/10.1016/j.envpol.2023.121857

3. Hussan H. N. M., He H., Du P., Wu X., Liu X., Xu J., Dong F., Zheng Y. Determination of clomazone and acetochlor residues in soybean (Glycine max (L.) Merr.). International Journal of Environmental Analytical Chemistry. 2021;101(15):2503–2509. https://doi.org/10.1080/03067319.2019.1694668

4. Li L., Yin Y., Zheng G., Liu S., Zhao C., Xie W., Ma L., Shan Q., Dai X., Wei L. Determination of multiclass herbicides in sediments and aquatic products using QuECHERS combined with ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and its application to risk assessment of rice-fish co-culture system in China. Microchemical Journal. 2021;170:106628. https://doi.org/10.1016/j.microc.2021.106628

5. Cara I. G., Topa D., Raus L., Calistru A. E., Filipov F., Jitareanu G. Selective and sensitive quantification of acetochlor and S-metolachlor in maize and soybean plant samples by gas chromatography-tandem mass spectrometry. Agriculture. 2021;11(4):283. https://doi.org/10.3390/agriculture11040283

6. Hegedűs G., Krikunova V., Bélai I., Eremin S., Székács A. An enzyme-linked immunosorbent assay (Elisa) for the detection of acetochlor. International Journal of Environmental Analytical Chemistry. 2002;82(11–12):879–891. https://doi.org/10.1080/0306731021000102338

7. Zha Y., Liu X., Hu P., Lu S., Ren H., Liu Z., Liang X., Yang Y., Li Y., Zhou Y. Alkaline phosphatase–triggered immunoassay based on fluorogenic reaction for sensitive detection of acetochlor, metolachlor, and propisochlor. Food Analytical Methods. 2020;13(4):1008–1016. https://doi.org/10.1007/s12161-020-01706-6

8. Li Z., Wu Y., Li Z., Yu B., Mao X., Shi G. A lateral flow immunoassay method for the rapid detection of acetochlor and alachlor in vegetable oil by sensitivity enhancement by using dimethyl-β-cyclodextrin. Analytical Methods. 2023;15(38):5087–5094. https://doi.org/10.1039/D3AY01379K

9. Yakovleva J. N., Lobanova A. I., Panchenko O. A., Eremin S A. Production of antibodies and development of specific polarization fluoroimmunoassay for acetochlor. International Journal of Environmental Analytical Chemistry. 2002;82(11–12):851–863. https://doi.org/10.1080/0306731021000102310

10. Altiparmak E., Yilmaz E., Dadaser-Celik F., Ates N. Sensitive quantification of acetochlor and metolachlor in water using Taguchi-optimized DLLME coupled with high-performance liquid chromatography. Microchemical Journal. 2024;200:110499. https://doi.org/10.1016/j.microc.2024.110499

11. Cai Y., Li L., Zhang J., Li Z., Zhang F., Xu Y., Tai Z. Development of a MOF-based SPE method combined with GC–MS for simultaneous determination of alachlor, acetochlor and pretilachlor in field soil. Environmental Monitoring and Assessment. 2023;195(5):569. https://doi.org/10.1007/s10661-023-11147-3

12. Ruan X., Wang Y., Kwon E. Y., Wang L., Cheng N., Niu X., Ding S., Van Wie B. J., Lin Y., Du D. Nanomaterial-enhanced 3D-printed sensor platform for simultaneous detection of atrazine and acetochlor. Biosensors and Bioelectronics. 2021;184:113238. https://doi.org/10.1016/j.bios.2021.113238

13. Liu Y., Wang L.-T., Zhou K., Eremin S. A., Huang X.-A., Sun Y.-M., Xu Z.-L., Lei H.-T. Rapid and homologous immunoassay for the detection of herbicide propisochlor in water. Food and Agricultural Immunology. 2018;29(1):67–83. https://doi.org/10.1080/09540105.2017.1359499

14. Mukhametova L. I., Eremin S. A. Fluorescence polarization assays for organic compounds in food safety. Frontiers in Bioscience-Elite. 2024;16(1):4. https://doi.org/10.31083/j.fbe1601004

15. Mukhametova L. I., Karimova M. R., Arutiunian D. A., Shanin I. A., Ponomarenko E. E., Eremin S. A. Detection of monobutyl phthalate in urine and environmental water by a rapid and sensitive fluorescence polarization immunoassay. One Health Advances. 2025;3(1):9. https://doi.org/10.1186/s44280-025-00074-2

16. Filimonova S. M., Balyklova K. S., Zherdev D. O., Eremin S. A., Mukhametova L. I., Krylov V. B., Nifantiev N. E. A First Case of Fluorescence Polarization Biosensor-Based Assay for Rapid Monitoring of Protein API Content in Tablet Dosage Forms: Detection of Lysozyme in Tablets. Biosensors. 2025;15(11):724. https://doi.org/10.3390/bios15110724

17. Deryabina M. A., Yakovleva Yu. N., Popova V. A., Eremin S. A. Determination of the herbicide acetochlor by fluorescence polarization immunoassay. Journal of Analytical Chemistry. 2005;60(1):80–85. https://doi.org/10.1007/s10809-005-0063-4