Impedimetric Polyaniline-Based Aptasensor for Aflatoxin B1 Determination in Agricultural Products

Abstract

An impedimetric aptasensor based on a polyaniline (PAni) support matrix is developed through the surface modification of a screen-printed carbon electrode (SPE) for aflatoxin B₁ (AFB₁) detection in foodstuffs and feedstuffs for food safety. The PAni is synthesized with the chemical oxidation method and characterized with potentiostat/galvanostat, FTIR, and UV-vis spectroscopy techniques. The stepwise fabrication procedure of the PAni-based aptasensor is characterized by means of cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) methods. The impedimetric aptasensor is optimized using the EIS technique, and its feasibility of detecting AFB₁ in real sample matrices is evaluated via a recovery study in spiked foodstuffs and feedstuffs, such as pistachio nuts, cinnamons, cloves, corn, and soybeans, with a good recovery percentage, ranging from 87.9% to 94.7%. The charge transfer resistance (R_CT) at the aptasensor interface increases linearly with the AFB₁ concentration in the range of 3 × 10^-2 nM to 8 × 10^-2 nM, with a regression coefficient (R²) value of 0.9991 and detection limit of 0.01 nM. The proposed aptasensor is highly selective towards AFB₁ and partially selective to AFB₂ and ochratoxin A (OTA) due to their similar structures that differ only at the carbon-carbon double bond located at C₈ and C₉ and the large molecule size of OTA.

Keywords: aptasensor; electrochemical impedance spectroscopy; electrochemical sensor; mycotoxin; polyaniline.

Figure 1

Schematic illustration of the impedimetric PAni-based aptasensor for AFB₁ detection.

Figure 2

(a) FTIR spectrum of chemical oxidative polymerized PAni measured by Attenuated Total Reflectance Fourier Transform Infrared (Perkin Elmer model FTIR spectrum 100 spectrometer). (b) UV–vis spectrum of chemical oxidative polymerized PAni captured by UV–vis spectrophotometer (Varian Cary 50). The chemical structure of PAni in ES form is shown in the inset.

Figure 3

(a) Cyclic voltammograms of PAni-modified SPE in 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl at different scan rates of 50 mV s⁻¹, 100 mV s⁻¹, 150 mV s⁻¹, 200 mV s⁻¹, 250 mV s⁻¹, and 300 mV s⁻¹. (b) Plot of anodic and cathodic peak currents against the square root of scan rate for PAni-modified SPE. (c) Plot of anodic and cathodic peak potentials against the logarithm of scan rate for PAni-modified SPE.

Figure 4

(a) Cyclic voltammograms of the bare SPE, PAni-modified SPE, PAni/Apt-modified SPE, and PAni/Apt-modified SPE during 5 nM AFB₁ detection at a scan rate of 100 mV s⁻¹ in the presence of 5 mM K₃[Fe(CN)₆] redox species. (b) Nyquist plots of bare SPE, PAni-modified SPE, PAni/Apt-modified SPE, and PAni/Apt-modified SPE during 5 nM AFB₁ detection in 13 mL of 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl at 25 °C.

Figure 5

(a) Nyquist plots for bare SPE and SPE modified with 3 μL and 4 μL of PAni in 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl. (b) Nyquist plots of PAni/Apt SPE during the detection of varying AFB₁ concentration ranging from 0.03 nM to 0.25 nM in 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl at pH 7.5. (c) The dynamic range of the aptasensor. The inset is the linear calibration range between 0.03 nM and 0.25 nM AFB₁ in 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl at pH 7.5. (d) Bar graph of the PAni-based aptasensor during the detection of AFB₁ with various interference species, such as AFB₂, OTA, OTB, and ZEN, in 5 mM K₃[Fe(CN)₆] redox indicator containing 0.1 M KCl at pH 7.5. (e) Bar chart of the reversibility of the AFB₁ aptasensor during 0.05 nM AFB₁ detection (30 min); regeneration of the aptasensor is achieved by incubating the aptasensor in 50 μM unmodified AFB₁ Apt solution as the regeneration solution for 30 min. The error bars indicate ± standard deviation, n = 3.