Yarynka D. Optical biosensor systems based on biomimetic polymers and smartphones for food mycotoxins (aflatoxin B1 and zearalenone) detection

Yarynka D.V. Optical biosensor systems based on biomimetic polymers and smartphones for food mycotoxins (aflatoxin B1 and zearalenone) detection. – The manuscript. A dissertation submitted in fulfillment of the requirements for the degree of Doctor of Philosophy (091 Biology). – Institute of Molecular Biology and Genetics of the National Academy of Sciences of Ukraine, Kyiv, 2023. Annually excessive levels of mycotoxins in food products are detected worldwide. In addition to the significant economic losses associated with crop loss (about 25% of food products are lost every year due to mycotoxins contamination), mycotoxins are dangerous for human and animal health. According to the Food and Agricultural Organization of the United Nations (FAO), the most dangerous mycotoxins are, among others, aflatoxin B1 (AFB1) and zearalenone (ZON). Therefore, developing new approaches for monitoring and controlling these mycotoxins in food products is very important. Thus, the dissertation aims to create optical biosensor systems based on smartphones and biomimetic polymers in the form of molecularly imprinted polymer membranes to detect worldwide food toxins (AFB1 and ZON) in food products and animal feed. Biomimetic polymers in the form of molecularly imprinted polymer (MIP) membranes are proposed as sensitive elements of biosensor systems to determine AFB1 and ZON. The toxin from analysed sample, bound with the selective receptor sites formed in the MIPs' structure, can be detected after irradiation of the membranes with ultraviolet (UV) light of a specific wavelength. It will initiate the fluorescence of mycotoxins, the intensity of which will be proportional to the concentration of toxins in the sample. Fluorescent sensor signals generated by the biomimetic polymers in the form of membranes can be registered both with a standard laboratory fluorimeter and with smartphone camera using commercially available digital image analysis programs for smartphones (in particular, the Spotxel program for the Android 6+ operating system) for further analysis. Using a smartphone as an analyser will simplify the analyte determination procedure and can successfully replace expensive laboratory equipment outside laboratories. Thus, this method can be used outside the laboratory. Firstly, highly stable sensitive elements based on MIP membranes with artificial receptor sites for AFB1 and ZON recognition in MIPs’ structure were synthesised by the in situ radical polymerisation method and optimised. The best recognition of AFB1 was found for MIP membranes synthesised using acrylamide (AA) as a functional monomer from a monomer mixture with a 1:2 “dummy” template: AA ratio. It was established that the MIP membranes synthesised using the functional monomers 1-allylpiperazine (1-ALP) and ethylene glycol methacrylate phosphate (EGMF), with 1:4 and 1:2 “dummy” template: functional monomer ratio, respectively, demonstrated the best recognition properties in response to adding ZON. Also, the influence of the analysed sample conditions on the ability of the synthesised MIP membranes to recognise the target mycotoxins was investigated and optimised. For the first time, laboratory prototypes of biosensor systems based on biomimetic polymers and smartphones for the mycotoxins analysis were created, and their analytical characteristics were investigated. In addition, to improve the working characteristics of proposed biosensor systems for highly sensitive mycotoxins analysis, new approaches were developed based on the use of a fluorescent marker in competitive analysis, as well as silver nanoparticles and plasmon-enhanced fluorescence. Prototypes of fluorescent biosensor systems based on smartphones and MIP membranes were successfully tested to determine AFB1 and ZON in wheat and corn real flour samples. In cooperation with the SE “Ukrmetrteststandard” the metrological studies of the developed sensor systems were carried out and the proposed methods for the mycotoxins analysis were approved.