Electrochemical aptasensor of estrogenic endocrine disrupting chemicals (EEDCS) based on modified aptamers and polypyrrole nanowires/gold nanoparticles

Abstract

The detection of estrogenic endocrine disrupting chemicals (EEDCs) such as 17β- estradiol (E2) and 17α-ethinyl estradiol (EE2) is a paramount importance towards the initiative on environmental monitoring due to its ability to disrupt endocrine system and trigger various cancer genes both in animals and humans. The alternative approach on the incorporation of modified or truncated aptamers into the sensor fabrication of this study is based on the earlier findings, which has proven lower binding affinities of extremely long (75,76-mers) and short aptamer sequence (<39-mers) that are unstable for sensor application. Henceforth, shorter aptamer may provide closer proximity between the aptamer and the target (improved binding affinity). Hence, this research involves the development of two different electrochemical aptasensors that has incorporated modified aptamers on the microspheres which were coated on the electrode fabricated with a conductive material (polypyrrole (PPY)/gold nanoparticles (Au)). Electrochemical properties of the aptasensors were characterised and validation test of the electrochemical aptasensors performance was conducted. The first successfully developed PPY/PMAA-NBA sensor for E2 detection in water sample are composed of two split DNA aptamers (synthesised from 12-mer and 14-mer aptamer truncated from 75-mer DNA aptamer) and polypyrrole nanowires/poly (methacrylic acid-co-n butyl acrylate) microspheres. In the presence of E2, the two split DNA aptamers were bound to E2 and established “split12-E2-split14 complex”, acting as a bridge on the surface of electrode and resulted to the detection of E2 by differential pulse voltammetry (DPV). The bridge acts as physical barrier on the electrode surface and inhibited redox reaction of [Fe(CN)6] -3/-4 solution, causes current response to decline and a relative current response was calculated. The second successfully developed electrochemical aptasensor for EE2 detection, Au/P(NBA-NAS), are composed of modified DNA aptamers (synthesised from 38-mer and 39-mer from 76- mer DNA aptamer that can selectively bind to EE2) and gold nanoparticles/poly(n-butyl acrylate-co-N-acryloxysuccinimide) microspheres. The signal generated from aptamer EE2 interaction was monitored electrochemically using DPV in the presence of [Fe(CN)6] -3/-4 solution. The measured current declined due to the EE2 bound interference and inhibited redox reaction on the electrode surface. Under pH 7.5 of 0.1 M Tris-HCl (100 mM NaCl), the fabricated PPY/PMAA-NBA aptasensor has shown the ability to detect E2 concentration in real samples in the range from 1×10-12 M to 1×10-4 M (R2 = 0.9772) with a detection limit of 0.48×10-12 M as well as 97 to 104 % of recovery (R2 = 0.9999) demonstrating high correlation of the sensor to the high performance liquid chromatography (HPLC) method. Otherwise, Au/P(NBA-NAS) aptasensor has exhibited a decline current trend, which was proportional to the EE2 concentration with the linear range from 1×10-6 M to 1×10-12 M (R2 = 0.9774 and R2 = 0.9845), with a detection limit of 0.24×10-12 M and 0.17×10-12 M, and 97 to 102 % of recovery (i.e. R2 = 0.9979 and R2 = 0.9953) highly correlated to the HPLC method), for 38-mer and 39-mer DNA aptamers, respectively. Both EE2 and E2 aptasensors were also able to distinguish these compounds from other EDCs compound.