10.57647/jnsc.2025.1504.15

Pretreatment-Free Rapid Detection of Tiamulin in Egg and Milk Samples using Gold Nanoparticles Based on Positive List System Regulations

  1. Department of Biological Sciences and Bioengineering, Inha University, Incheon, Republic of Korea
  2. School of Chemical Engineering, Sungkyunkwan University, Suwon 16419, Republic of Korea
  3. National Institute of Animal Science, Rural Development Administration, Wanju, South Korea

Received: 27-05-2025

Revised: 12-08-2025

Accepted: 31-08-2025

Published in Issue 31-08-2025

How to Cite

Alhammadi, M., Sonwal, S., Han, S., Kim, H., Kim, D.-H., Oh, M.-H., & Huh, Y. S. (2025). Pretreatment-Free Rapid Detection of Tiamulin in Egg and Milk Samples using Gold Nanoparticles Based on Positive List System Regulations. Journal of Nanostructure in Chemistry, 15(4). https://doi.org/10.57647/jnsc.2025.1504.15

PDF views: 200

Abstract

Tiamulin (TML) antibiotic residues in animal products could pose potential health hazards to consumers. South Korea set the positive list system regulations, where the maximum residue limits of TML in poultry eggs should not exceed 1000 ng/mL. Therefore, we developed a simple and pretreatment-free lateral flow immunoassay based on a highly specific TML antibody conjugated with gold nanoparticles for qualitative and semi-quantitative analysis of TML. The optimized sensor showed a visual limit of detection of 15 ng/mL for both milk- and egg-spiked samples, with a cut-off value of 100 ng/mL and 180 ng/mL for milk and egg, respectively. The assay demonstrated high recovery in both milk and egg matrices with relative standard deviation values below 12%, confirming its suitability for rapid and reliable determination of TML in complex food samples. As both sample types required no pretreatment and provided visible results within 10 minutes, the sensor is simple, user-friendly, and highly tolerant to matrix interferences. Moreover, it exhibited specific selectivity toward TML, supporting its potential as a promising detection platform for monitoring TML in animal products.

Keywords

  • Immunochromatographic assay,
  • On-site detection,
  • Antibody,
  • Tiamulin,
  • Pretreatment-free,
  • Gold nanoparticles

References

  1. P. Cybulski, A. Gajda, M. Bilecka, and A. Jabłoński, Determination of Tiamulin Concentration in Sow Milk and in Sera of Suckling Piglets, Molecules 28, 6940 (2023).
  2. M. D. Borak, L. Sarc, D. G. Mugerli, B. Antolic, and M. Brvar, Occupational inhalation poisoning with the veterinary antibiotic tiamulin, Clin. Toxicol. 58, 287 (2020).
  3. E. Pankowska, O. Kończak, P. Żakowicz, T. Wojciechowicz, M. Gogulski, and L. Radko, Protective Action of Cannabidiol on Tiamulin Toxicity in Humans—In Vitro Study, Int. J. Mol. Sci. 25, 13542 (2024).
  4. J. Oszmianski, S. Lachowicz, W. Wi´sniewska, V. C. Ciucă, C. O. Rusănescu, and V. V. Safta, Analysis of Transfer of Tiamulin to Animal Tissue after Oral Administration: An Important Factor for Ensuring Food Safety and Environmental Protection, Pharmaceuticals 16, 387 (2023).
  5. Ministry of Food and Drug Safety, https://www.mfds.go.kr/eng/index.do.
  6. E. Patyra, C. Nebot, R. E. Gavilán, A. Cepeda, and K. Kwiatek, Development and validation of an LC-MS/MS method for the quantification of tiamulin, trimethoprim, tylosin, sulfadiazine and sulfamethazine in medicated feed, Food Addit. Contam.: Part A. 35, 882 (2018).
  7. F. Sun et al., Comprehensive analysis of tiamulin metabolites in various species of farm animals using ultra-high-performance liquid chromatography coupled to quadrupole/time-of-flight, J. Agric. Food Chem. 65, 199 (2017).
  8. X. You et al., A novel electrochemical immunosensor for the sensitive detection of tiamulin based on staphylococcal protein A and silver nanoparticle-graphene oxide nanocomposites, Bioelectrochemistry 141, 107877 (2021).
  9. J. Zhou, X. Zhang, W. Qian, Q. Yang, Y. Qi, Y. Chen, and A. Wang, Quantum dots-based fluorescence immunoassay for detection of tiamulin in pork, J. Food Saf. 41, e12930 (2021).
  10. L. Li, J. Y. Yang, Y. Wang, Z. Zhang, T. T. Yuan, Y. M. Xiao, Z. L. Xu, and Y. D. Shen, Development of a Chemiluminescence Enzyme Immunoassay Method for Detection of Tiamulin Residues in Eggs, J. Instrum. Anal. 41, 115 (2022).
  11. M. F. Lanjwani, N. Altunay, and M. Tuzen, Preparation of fatty acid-based ternary deep eutectic solvents: Application for determination of tetracycline residue in water, honey and milk samples by using vortex-assisted microextraction, Food Chem. 400, 134085 (2023).
  12. M. Nemati, M. R. Afshar Mogaddam, M. A. Farazajdeh, M. Tuzen, and J. Khandaghi, In-situ formation/decomposition of deep eutectic solvent during solidification of floating organic droplet-liquid-liquid microextraction method for the extraction of some antibiotics from honey prior to high performance liquid chromatography-tandem mass spectrometry, J. Chromatogr. A 1660, 462653 (2021).
  13. A. C. Mirica, D. Stan, I. C. Chelcea, C. M. Mihailescu, A. Ofiteru, and L. A. Bocancia-Mateescu, Latest Trends in Lateral Flow Immunoassay (LFIA) Detection Labels and Conjugation Process, Front Bioeng. Biotechnol. 10, 922772 (2022).
  14. K. H. Kim et al., Low-Powered pH-Stable Nano-electrokinetically Enhanced Lateral Flow Assay for COVID-19 Antigen Test, Biochip J. 17, 340 (2023).
  15. F. Zhao, K. Wangpimool, and J. C. Kim, Near-infrared and Thermo-sensitive Liposomes Incorporating Thiolated-carboxymethyl Cellulose-capped Gold Nanoparticles and Poly(N-isopropylacrylamide), Biotechnol. Bioprocess Eng. 28, 589 (2023).
  16. Q. Dai, S. Tang, and C. Dai, Recent Advances in Pretreatment Methods and Detection Techniques for Veterinary Drug Residues in Animal-Derived Foods, Metabolites 15, 233 (2025).
  17. X. Lei, S. Song, H. Tao, L. Liu, Q. Zheng, C. Xu, and H. Kuang, Development of Indirect Competitive Enzyme-Linked Immunosorbent and Immunochromatographic Strip Assays for Tiamulin Detection in Chicken, ACS Omega 3, 3581 (2018).
  18. G. Na, X. Hu, Y. Sun, S. Kwee, G. Xing, Y. Xing, and G. Zhang, A highly sensitive monoclonal antibody−based paper sensor for simultaneously detecting valnemulin and tiamulin in porcine liver, J. Food Sci. 85, 1681 (2020).
  19. J. Dong, P. L. Carpinone, G. Pyrgiotakis, P. Demokritou, and B. M. Moudgil, Synthesis of Precision Gold Nanoparticles Using Turkevich Method, Kona 37, 224 (2020).
  20. M. Alhammadi, S. Aliya, R. Umapathi, M. H. Oh, and Y. S. Huh, A simultaneous qualitative and quantitative lateral flow immunoassay for on-site and rapid detection of streptomycin in pig blood serum and urine, Microchem. J. 195, 109427 (2023).
  21. Y. Moon, H. H. Cho, H. Moon, H. Song, J. C. Ro, J. H. Lee, and J. Lee, Simultaneous Triplex Detection in a Single-Test-Line Lateral Flow Immunoassay Utilizing Distinct Nanoparticle Colorimetry, Biochip J. 18, 247 (2024).
  22. G. A. Posthuma-Trumpie, J. Korf, and A. Van Amerongen, Lateral flow (immuno)assay: Its strengths, weaknesses, opportunities and threats. A literature survey, Anal. Bioanal. Chem. 393, 569 (2009).
  23. V. P. Shah et al., Bioanalytical method validation - A revisit with a decade of progress, Pharm. Res. 17, 1551 (2000).
  24. H. Tyagi, A. Kushwaha, A. Kumar, and M. Aslam, A Facile pH Controlled Citrate-Based Reduction Method for Gold Nanoparticle Synthesis at Room Temperature, Nanoscale. Res. Lett. 11, 1 (2016).
  25. D. Philip, Green synthesis of gold and silver nanoparticles using Hibiscus rosa sinensis, Physica E Low Dimens. Syst. Nanostruct. 42, 1417 (2010).
  26. J. O. Tam, H. de Puig, C. wan Yen, I. Bosch, J. Gómez-Márquez, C. Clavet, K. Hamad-Schifferli, and L. Gehrke, A Comparison of Nanoparticle-Antibody Conjugation Strategies in Sandwich Immunoassays, J. Immunoassay Immunochem. 38, 355 (2016).
  27. J. P. Oliveira, A. R. Prado, W. J. Keijok, P. W. P. Antunes, E. R. Yapuchura, and M. C. C. Guimarães, Impact of conjugation strategies for targeting of antibodies in gold nanoparticles for ultrasensitive detection of 17β-estradiol, Sci. Rep. 9, 1 (2019).
  28. O. D. Hendrickson, N. A. Byzova, E. A. Zvereva, A. V. Zherdev, and B. B. Dzantiev, Sensitive lateral flow immunoassay of an antibiotic neomycin in foodstuffs, J. Food Sci. Technol. 58, 292 (2021).
  29. Y. Wang, B. Ma, M. Liu, E. Chen, Y. Xu, and M. Zhang, Europium Fluorescent Nanoparticles-Based Multiplex Lateral Flow Immunoassay for Simultaneous Detection of Three Antibiotic Families Residue, Front. Chem. 9, 793355 (2021).
  30. M. Alhammadi, J. Yoo, S. Sonwal, S. Y. Park, R. Umapathi, M. H. Oh, and Y. S. Huh, A highly sensitive lateral flow immunoassay for the rapid and on-site detection of enrofloxacin in milk, Front. Nutr. 9, 1036826 (2022).
  31. H.-C. ; Chen, S.-H. ; Cheng, Y.-H. ; Tsai, D.-F. Hwang, H.-C. Chen, S.-H. Cheng, Y.-H. Tsai, and A. D.-F. Hwang, Determination of tiamulin residue in pork and chicken by solid phase extraction and HPLC, J. Food Drug Anal. 14, 2 (2020).
  32. A. Wang et al., Development of a label free electrochemical sensor based on a sensitive monoclonal antibody for the detection of tiamulin, Food Chem. 366, 130573 (2022).