Development of electrochemical biosensors for sweeteners using engineered nanomaterials supported by computational modelling

Abstract

Electrochemical immunosensors are a powerful tool in analytical applications. The current methods for the isolation and detection of artificial and natural sweeteners suffer from challenges in sample preparation and a lack of specificity. However, electrochemical immunosensors offer a sensitive, economical, and selective analytical solution to analyse these commonly used sweeteners, such as aspartame. The author of this work developed electrochemical immunosensors for the food and beverage industries to use in the detection and measurement of aspartame, a non-nutritive sweetener, and rebaudioside A, a natural sweetener. Most artificial sweeteners are low-calorie options that are suggested for ailments linked to health. These sweeteners' ability to remain stable at even high temperatures has greatly expanded the range of meals that can use them. Aspartame and rebaudioside A have not been linked to any health risks, although regulation is still necessary because of their extensive use in the food industry. The developed immunosensors for the detection of aspartame and rebaudioside A were achieved and presented as three case studies in this study. In the first case study, the immunosensor was achieved by fabricating green synthesized PVP capped silver nanoparticles (PVP-AgNPs) with functionalized multi-walled carbon nanotubes (fMWCNTs) and immobilizing the human sweet tase receptor T1R2 in a glassy carbon electrode (GCE), resulting in GCE/PVP-AgNPs/fMWCNTs/T1R2. The electrochemical assessment of aspartame was achieved using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and differential pulse voltammetry (DPV), respectively, under optimum pH 8 in a 0.1 M phosphate buffer with reference to the Ag/AgCl reference electrode. The electro-oxidation of ASP was noticed by a well-defined oxidation peak potential at 1.4 V. The immunosensor sensor showed a linear dynamic range of 2.89 to 27.61 μM (R 2 = 0.9170) based on differential pulse voltammetry, with limits of detection (LOD) and quantification (LOQ) (S/N = 3) of 0.40 μM and 1.34 μM, respectively. The second case study focused on the indirect electrochemical detection of rebaudioside A in the presence of ferro/ferricyanide as a redox probe. The immunosensor was developed by fabricating GCE with zeolitic imidazolate framework-67 (ZIF-67) in combination with fMWCNTs and the immobilization of the T1R2 receptor. The qualitative and quantitative analysis of rebaudioside A was done using CV, EIS, and DPV utilizing a 5 mM [Fe (CN)6] 3-/-4 redox probe. The stable electrode had an exponential dynamic range of 0.9901 µM to 8.2569 µM (R2 = 0.9996). The LOD and LOQ were computed to be 1.10 µM and 3.33 µM, respectively. This case study also used Patch Dock and PyRx to better understand the interactions between Reb A and T1R2. The final case study employed a platinum electrode (PtE) as the working electrode (WE) for the electrochemical immunosensing of aspartame. The modification of PtE involved the utilization of a nanocomposite consisting of PVP-AgNPs and reduced graphene oxide (rGO), with T1R2 immobilized. The electrochemical detection of aspartame was achieved under optimized conditions at pH 8 in a 0.1 M phosphate buffer, utilizing CV, EIS, and DPV as electrochemical tools. The PVP-AgNPs/rGO/T1R2 was used to fabricate Pt and the electrode performed well with a linear increase in oxidation peak currents as aspartame concentrations were increased from 2.38 µM to 25.78 µM (0.9529). The LOD and LOQ were calculated to be 5.85 µM and 17.73 µM, respectively. The synthesized nanoparticles and nanocomposites (PVP-AgNPs/fMWCNTs, ZIF 67/fMWCNTs, and PVP-AgNPs/rGO) were characterized using conventional techniques such as UV-Vis spectroscopy, thermogravimetric analysis (TGA), Fourier transform infrared spectroscopy (FTIR), field flow fractionation (FFF), single particle inductively coupled plasma mass spectrometry (sp-ICPMS), energy-dispersive X-ray spectroscopy (EDS), and scanning electron microscopy (SEM). In addition to the experimental results, computational chemistry methods were undertaken. These included adsorption assessments, density functional theory (DFT), and molecular docking techniques. These techniques were all aimed at achieving a deeper molecular-level understanding of the interactions among the analytes (Aspartame and Reb A), T1R2, and the nanocomposites employed in the modification of the working electrodes (GCE and Pt-E)