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Antibody-free colorimetric determination of total aflatoxins by mercury(II)-mediated aggregation of lysine-functionalized gold nanoparticles


Currently known immunoassays for aflatoxins (AFs) usually are not capable of simultaneous determination of total AFs (i.e., aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2) due to the lack of group-specific antibodies. We are presenting here a colorimetric method for determination of total AFs by exploiting the Hg(II) ion-mediated aggregation of lysine-functionalized gold nanoparticles (Lys-AuNPs). AFs bind Hg(II) ion to form stable complexes, and this prevents the Hg(II) induced aggregation of Lys-AuNPs. Hence, the color change of solutions of AuNPs that occurs after aggregation of the AuNPs is suppressed. This effect was exploited to design a method for quantitation of AFs that can be detected by UV-Vis spectrophotometry (by measuring the ratio of absorbances at 725 nm and 525 nm), and even with bare eyes. The method has a very low detection limit (1.1 ppb) and is highly selective for AFs over other mycotoxins. Colorimetric analysis of rice samples by this method gave results that were in good agreement with those obtained by HPLC. To the best of our knowledge, this is the first method for visual detection of total AFs based on the distance-dependent optical properties of AuNPs.

Aflatoxins (AFs) coordinate with Hg2+ ions to form AF-Hg2+ complexes and prevent the Hg2+-induced aggregation of lysine-modified gold nanoparticles (Lys-AuNPs). This results in a gray-blue to red color change of solutions and enables the determination of total AFs by UV-Vis spectrophotometry or bare eyes.

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  1. 1.

    Shephard GS (2008) Determination of mycotoxins in human foods. Chem Soc Rev 37:2468–2477

    CAS  Article  Google Scholar 

  2. 2.

    International Agency for Research on Cancer (2002) IARC monographs on the evaluation of carcinogenic risks to humans. IARC 82:171–300

    Google Scholar 

  3. 3.

    U. S. Food and Drug Administration (1994) Compliance policy guide sec. 683.100 - action levels for aflatoxins in animal feeds.

  4. 4.

    Administration of Quality Supervision, Inspection and Quarantine of China (2011) GB 2761-2011, maximum levels of mycotoxins in foods.

  5. 5.

    Golge O, Hepsag F, Kabak B (2016) Determination of aflatoxins in walnut sujuk and Turkish delight by HPLC-FLD method. Food Control 59:731–736

    CAS  Article  Google Scholar 

  6. 6.

    Campone L, Piccinelli AL, Celano R, Russo M, Valdés A, Ibáñez C, Rastrelli L (2015) A fully automated method for simultaneous determination of aflatoxins and ochratoxin A in dried fruits by pressurized liquid extraction and online solid-phase extraction cleanup coupled to ultra-high-pressure liquid chromatography-tandem mass spectrometry. Anal Bioanal Chem 407:2899–2911

    CAS  Article  Google Scholar 

  7. 7.

    Zhang H, Mo HZ, Li H (2013) Preparation of aflatoxin B1 polyclonal antibody and development of an indirect competitive enzyme-linked immunosorbent assay. J Food Agric Environ 11:190–194

    CAS  Google Scholar 

  8. 8.

    Yu FY, Gribas AV, Vdovenko MM, Sakharov IY (2013) Development of ultrasensitive direct chemiluminescent enzyme immunoassay for determination of aflatoxin B1 in food products. Talanta 107:25–29

    CAS  Article  Google Scholar 

  9. 9.

    Delmulle BS, De Saeger SMDG, Sibanda L, Barna-Vetro I, Van Peteghem CH (2005) Development of an immunoassay-based lateral flow dipstick for the rapid detection of aflatoxin B1 in pig feed. J Agric Food Chem 53:3364–3368

    CAS  Article  Google Scholar 

  10. 10.

    Tang DP, Liu BQ, Niessner R, Li PW, Knopp D (2013) Target-induced displacement reaction accompanying cargo release from magnetic mesoporous silica nanocontainers for fluorescence immunoassay. Anal Chem 85:10589–10596

    CAS  Article  Google Scholar 

  11. 11.

    Wang Y, Liu N, Ning BA, Liu M, Lv Z, Sun ZY, Peng Y, Chen CC, Li JW, Gao ZX (2012) Simultaneous and rapid detection of six different mycotoxins using an immunochip. Biosens Bioelectron 34:44–50

    Article  Google Scholar 

  12. 12.

    He QH, Xu Y, Wang D, Kang M, Huang ZB, Li YP (2012) Simultaneous multiresidue determination of mycotoxins in cereal samples by polyvinylidene fluoride membrane based dot immunoassay. Food Chem 134:507–512

    CAS  Article  Google Scholar 

  13. 13.

    Xu GF, Zhang SP, Zhang QR, Gong LS, Dai H, Lin YY (2016) Magnetic functionalized electrospun nanofibers for magnetically controlled ultrasensitive label-free electrochemiluminescent immunedetection of aflatoxin B1. Sensors Actuators B Chem 222:707–713

    CAS  Article  Google Scholar 

  14. 14.

    Wang YW, Li PW, Majkova Z, Bever CRS, Kim HJ, Zhang Q, Dechant JE, Gee SJ, Hammock BD (2013) Isolation of alpaca anti-idiotypic heavy-chain single-domain antibody for the aflatoxin immunoassay. Anal Chem 85:8298–8303

    CAS  Article  Google Scholar 

  15. 15.

    Lee KM, Herrman TJ, Bisrat Y, Murray SC (2015) Feasibility of surface-enhanced Raman spectroscopy for rapid detection of aflatoxins in maize. J Agric Food Chem 62:4466–4474

    Article  Google Scholar 

  16. 16.

    Zhang Q, Jia FG, Liu CH, Sun JK, Zheng XZ (2014) Rapid detection of aflatoxin B1 in paddy rice as analytical quality assessment by near infrared spectroscopy. Inter J Agr Biol Eng 7:127–133

    Google Scholar 

  17. 17.

    Wang W, Ni XZ, Lawrence KC, Yoon SC, Heitschmidt GW, Feldner P (2015) Feasibility of detecting Aflatoxin B1 in single maize kernels using hyperspectral imaging. J Food Eng 166:182–192

    CAS  Article  Google Scholar 

  18. 18.

    Xiao C, Liu JF, Yang AK, Zhao H, He YJ, Li XJ, Yuan ZB (2015) Colorimetric determination of neomycin using melamine modified gold nanoparticles. Microchim Acta 182:1501–1507

    CAS  Article  Google Scholar 

  19. 19.

    Jiang Y, Zhao H, Zhu NN, Lin YQ, Yu P, Mao LQ (2008) A simple assay for direct colorimetric visualization of trinitrotoluene at picomolar levels using gold nanoparticles. Angew Chem Int Ed 47:8601–8604

    CAS  Article  Google Scholar 

  20. 20.

    Ai KL, Liu YL, Lu LH (2009) Hydrogen-bonding recognition-induced color change of gold nanoparticles for visual detection of melamine in raw milk and infant formula. J Am Chem Soc 131:9496–9497

    CAS  Article  Google Scholar 

  21. 21.

    Xu JY, Li Y, Bie JX, Jiang W, Guo JJ, Luo YL, Shen F, Sun CY (2015) Colorimetric method for determination of bisphenol A based on aptamer-mediated aggregation of positively charged gold nanoparticles. Microchim Acta 182:2131–2138

    CAS  Article  Google Scholar 

  22. 22.

    Xu H, Wang YW, Huang XM, Li Y, Zhang H, Zhong XH (2012) Hg2+-mediated aggregation of gold nanoparticles for colorimetric screening of biothiols. Analyst 137:924–931

    CAS  Article  Google Scholar 

  23. 23.

    Song J, Huang PC, Wan YQ, Wu FY (2016) Colorimetric detection of thiocyanate based on anti-aggregation of gold nanoparticles in the presence of cetyltrimethyl ammonium bromide. Sensors Actuators B Chem 222:790–796

    CAS  Article  Google Scholar 

  24. 24.

    Zhang M, Zhang YH, Ma L (2011) Studies and application of fluorescence of aflatoxin B1 enhanced by synergetic effect of β-cyclodextin and its derivatives and metalions. Chin J Anal Chem 39:907–1911

    Google Scholar 

  25. 25.

    Chen ZB, Zhang CM, Tan Y, Zhou TH, Ma H, Wan CQ, Lin YQ, Li K (2015) Chitosan-functionalized gold nanoparticles for colorimetric detection of mercury ions based on chelation-induced aggregation. Microchim Acta 182:611–616

    CAS  Article  Google Scholar 

  26. 26.

    Sener G, Uzun L, Denizli A (2014) Lysine-promoted colorimetric response of gold nanoparticles: A simple assay for ultrasensitive mercury(II) detection. Anal Chem 86:514–520

    CAS  Article  Google Scholar 

  27. 27.

    Administration of Quality Supervision, Inspection and Quarantine of China (2003) GB/T 5009.22-2003, Determination Aflatoxin B1 in Foods.

  28. 28.

    Qi PP, Wang ZW, Yang GL, Shang CQ, Xu H, Wang XY, Zhang H, Wang Q, Wang XQ (2015) Removal of acidic interferences in multi-pesticides residue analysis of fruits using modified magnetic nanoparticles prior to determination via ultra-HPLC-MS/MS. Microchim Acta 182:2521–2528

    CAS  Article  Google Scholar 

  29. 29.

    Haiss W, Thanh TKN, Aveyard J, Fernig DG (2007) Determination of size and concentration of gold nanoparticles from UV-Vis spectra. Anal Chem 79:4215–4221

    CAS  Article  Google Scholar 

  30. 30.

    Shim WB, Kim MJ, Mun H, Kim MG (2014) An aptamer-based dipstick assay for the rapid and simple detection of aflatoxin B1. Biosens Bioelectron 62:288–294

    CAS  Article  Google Scholar 

  31. 31.

    Wang D, Hu WH, Xiong XH, Xu Y, Li CM (2015) Multifunctionalized reduced graphene oxide-doped polypyrrole/pyrrolepropylic acid nanocomposite impedimetric immunosensor to ultra-sensitively detect small molecular aflatoxin B1. Biosens Bioelectron 63:185–189

    CAS  Article  Google Scholar 

  32. 32.

    Liu ZX, Gao JX, Yu JJ (2006) Aflatoxins in stored maize and rice grains in Liaoning province, China. J Stored Prod Res 42:468–479

    CAS  Article  Google Scholar 

  33. 33.

    Firdous S, Ashfaq A, Khan SJ, Khan N (2014) Aflatoxins in corn and rice sold in Lahore, Pakistan. Food Addit Contam B 7:95–98

    CAS  Article  Google Scholar 

  34. 34.

    Administration of Quality Supervision, Inspection and Quarantine of China (2003) GB/T 18979-2003. Content in Food, Determination Aflatoxins

    Google Scholar 

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This work was sponsored by the National Natural Science Foundation of China (Grant No. 21272263, 31201832), the State Key Laboratory of Natural and Biomimetic Drugs (No. K20140204), and the National S&T Pillar Project (2011BAD26B0405). We thank Yang Li (Chinese Academy of Agricultural Sciences) for the help of mass spectrometry and Chenchen Guo (University of Chinese Academy of Sciences) for the help of theoretical calculation.

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Correspondence to Jingkui Yang or Yujian He.

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The author(s) declare that they have no competing interests

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Bibai Du and Peilong Wang contributed equally to this work

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Du, B., Wang, P., Xiao, C. et al. Antibody-free colorimetric determination of total aflatoxins by mercury(II)-mediated aggregation of lysine-functionalized gold nanoparticles. Microchim Acta 183, 1493–1500 (2016).

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  • Spectrophotometry
  • Group specific assay
  • Visual test
  • Colloid
  • Transmission electron microscopy
  • Dynamic light scattering
  • Density functional theory
  • HPLC
  • Food analysis