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Start date: 2021Subject: search.filters.subject.Mycotoxins

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    Development of electrochemical sensors for the detection of mycotoxins in food matrices using functionalised nanocomposites
    (2024-05) Naidoo, Lyndon; Bisetty, Krishna; Meier, Florian; Uwaya, Gloria Ebube
    The analysis of pathogens in foods is of critical importance to ensure consumer safety and quality assurance, as contaminants pose serious risks to public health. Mycotoxins are naturally occurring carcinogenic toxins that arise from specific strains of fungi as they contaminate food. They are found in a wide variety of grains, cereals, and dairy products, causing cancer in both humans and animals. Thus, there is a growing demand for simple, sensitive and inexpensive sensors for mycotoxin detection in lieu of conventionally employed large-scale instrumentation. In this study, the development of electrochemical sensors for the detection of aflatoxin B1 (AFB1), zearalenone (ZEN) and ochratoxin A (OTA) in foods was investigated and presented as three case studies, respectively. In the first case study, an ultrasensitive aptasensor was developed for the indirect detection of AFB1 in the presence of a ferri/ferrocyanide ([Fe(CN)6]3-/4-) redox probe solution. The sensor was constructed by immobilizing an anti-AFB1 aptamer (Apt) to a carboxylated multiwalled carbon nanotube (cMWCNT) and iron oxide (Fe3O4) nanoparticle (NP) composite using a glassy carbon electrode (GCE). This resulted in the development of the GCE/cMWCNTsFe3O4 NP/Apt sensor. An electrochemical response was exhibited from AFB1 applying cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and differential pulse voltammetry (DPV), respectively, utilizing a [Fe(CN)6]3-/4- redox probe prepared in phosphatebuffered saline (PBS) solution with reference to the Ag/AgCl reference electrode under optimized conditions. DPV findings reported very low limits of detection (LOD) and quantification (LOQ) of 0.43 fg mL-1 and 1.44 fg mL-1 respectively in comparison to current literature, over a calibration range of 0.50 fg mL-1 to 5.00 fg mL-1. For real sample analysis, excellent spike recoveries from 95% to 105% were obtained for corn and rice flour. Density functional theory (DFT) was used to propose a reaction scheme by ascertaining the electronic properties of the redox-active functional groups of AFB1. This supported the experimental anodic response findings of DPV. The second case study focused on how PEGylated Fe3O4 NPs and cMWCNTs fabricated on a GCE could be used for the design of an electrochemical sensor for ZEN analysis. The qualitative and quantitative analyses of ZEN were completed using CV, EIS and DPV, respectively, under optimized conditions in a sodium phosphate buffer solution. The developed sensor reported significantly low LODs and LOQs of 0.34 fg mL-1 and 1.12 fg mL-1 respectively, over a calibration range of 1.00 fg mL-1 to 10.00 fg mL-1 by DPV. Excellent spike recoveries ranging from 92% to 106% were obtained for rice and corn flour. The Monte Carlo (MC) adsorption simulation studies predicted the strong interaction of ZEN with the constructed sensor. In the final case study, an OTA electrochemical sensor was designed using a nickel metalorganic framework (Ni-MOF) and carboxylated reduced graphene oxide (cRGO) on a GCE. The detection of OTA was achieved under optimized conditions in PBS solution with the developed GCE/Ni-MOF/cRGO electrode, employing CV, EIS and DPV as electrochemical tools. Applying DPV, the sensor reported very low LODs and LOQs of 3.29 fg mL-1 and 10.97 fg mL-1 respectively, over a calibration range of 10.00 fg mL-1 to 90.00 fg mL-1. Regarding real sample analysis, excellent spike recoveries from 95% to 105% were obtained for corn and rice flour. Molecular dynamics (MD) studies predicted that the Ni-MOF exhibited a strong electrostatic interaction with the OTA analyte, in agreement with the experimental findings. The synthesized nanomaterials (cMWCNTs-Fe3O4 NP, PEG-Fe3O4 NPs/cMWCNTs, and NiMOF/cRGO) utilized in this study were characterized by an array of techniques, including single particle inductively coupled plasma-mass spectrometry (spICP-MS), transmission electron microscopy (TEM), X-ray diffraction (XRD), energy-dispersive X-ray spectroscopy (EDS), scanning electron microscopy (SEM), multidetector asymmetrical flow field-flow fractionation (AF4), and Fourier transform infrared spectroscopy (FTIR). Finally, computational modelling studies were undertaken to establish a synergy with the experimental approaches employed in each case study. These methodologies included DFT, docking studies, MC adsorption and MD simulations, which were aimed at predicting and assessing the atomic and molecular interactions between the mycotoxins and their respective electrode systems.
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    Electrochemical aptasensor for the detection of mycotoxins in food samples by experimental and computational methods
    (2021) Kunene, Kwanele; Bisetty, Krishna; Kanchi, Suvardhan; Sabela, Myalowenkosi Innocent
    Mycotoxins are secondary metabolites of fungi that are present in various foodstuff and feed commodities. A large number of mycotoxins exist, however only a limited number represent a major damages and toxic properties. Amongst them, the aflatoxins and ochratoxins are deemed to be the most poisonous and extensively circulated in the world and then, represent a real hazard to both human and animal. Depending on several factors like the consumption levels, exposure time, mechanisms of action, digestion and defense mechanisms, mycotoxins stimulate a wide spectrum of toxicological effects leading to both acute and chronic diseases, liver and kidney failure, skin rash, cancer, immune suppression, birth defects or even death. To address the harmful impact of mycotoxin contaminants in food and feed, health authorities in various countries world-wide have established guidelines in order to protect human and animal from the possible damages instigated by these toxins. Authorities such as the European Commission, US Food and Drug Administration (FDA), World Health Organization (WHO) and the Food and Agriculture Organization of the United Nations (FAO) set up maximum level regulations for main mycotoxins in foods and feeds. To accomplish the expectations of these regulation levels, there is a great need for the development and validation of modern, uncomplicated, rapid, and detailed methodologies for the detection of toxins. In this study, various approaches for the rapid, inexpensive and ultrasensitive biosensors for the detection of two major mycotoxins were developed. The electrochemical-based aptasensor and immunosensor were developed for the determination of aflatoxin B1 (AFB1) and ochratoxin A (OTA) in different food products. The fabricated biosensors demonstrated good practical analytical feasibility for mycotoxins detection in real samples such as WeetBix, yoghurt, coffee and in wine samples with excellent recoveries and RSD values. To avoid fouling on the sensor surface by the constituents present in real samples, the carbon screen printed electrode (C-SPE) and carbon felt electrode (CFE) surfaces were modified with different nanomaterials such as silver nanoparticle (AgNPs), palladium nanoparticles (PdNPs), palladium doped boron nitride (PdNPs-BN) and titanium nanoparticles doped with boron nitride BN-TiO2. In addition, the aptamers and antibodies were immobilized on the modified electrode in order to enhance the selectivity of the sensor towards the detection of OTA and AFB1. The electrochemical aptasensor for OTA permitted for highly sensitive detection in Weet-Bix with a wide linear range (0.002 - 0.016 mg L-1) and limit of detection of 7×10-4 mg L-1. It is worth prominence that it is the first time that carbon screen printed electrode (C-SPE) modified with AgNPs was used, opening new pathways for highly precise analysis. Experimental results were further supported computationally for a better understanding of the interaction between the aptamer and the analytes. Computational results were in good agreement with experimental results. The same procedure was also established in voltammetric detection of AFB1 using CFE modified with BN-TiO2 (CF/BN-TiO2). A wide concentration range of 2.5 - 20 ng mL-1 with an excellent LOD of 0.002 ng mL-1 for AFB1 was obtained. For the case study of wine samples tested for AFB1 detection, a simple but very effective pretreatment method was effectively applied. The addition of acetonitrile to the wine reduces the non-specific interactions that might be accountable for inactivation of antibody and blocking of the sensor surface. Furthermore, the PdNPs-BN enhanced the electrical signal and the sensor sensitivity. Attained results allowed for AFB1 detection at concentrations range from 1.0 - 10 ng mL-1 with limit of detection of 0.832 ng mL-1 . In the case study of the electrochemical immunosensor for the detection of OTA in coffee, a linear detection range of 0.5 - 20 ng mL-1 was achieved with LOD of 0.096 ng mL-1 . The fabricated aptasensors and immunosensors in this study combines the most desirable characteristics of a good biosensor such as high sensitivity, inexpensive, rapid, and simple but portable method make proposed approaches an important and very promising tools for extensive biosensing applications.