Biosensors for Environmental Monitoring via Bioinhibition

摘要

Biosensors are devices that recognize, analyze, and transduce signal for detection via biological materials with bio-recognition properties. As of now, almost all kinds of biomaterials, including microbial cells, organelles, proteins, and nucleic acid, have been used for biosensors. In this dissertation, two kinds of biomaterials, enzymes and organelles, have been used for novel biosensor development. The primary focus of the research is the inhibition mechanism of laccase and mitochondria by different environmental toxins, like arsenic and pesticides.;In Chapter 2, a laccase-based biosensor was developed for arsenic sensing. Inhibition of oxygen reduction by arsenic was observed electrochemically via laccase immobilized with anthracene-modified multiwalled carbon nanotubes on a Toray carbon paper electrode. First, it was found that laccase is inhibited by arsenite and arsenate, and the inhibition mechanism was further determined as mixed inhibition (with preference to an uncompetitive inhibition model). Second, the laccase-modified electrodes were then fabricated into a self-powered biosensor with flavin-adenine-dinucleotide-dependent glucose dehydrogenase-based bioanodes. The biosensor was operated at 10% of its maximum current and demonstrated a detection limit of 13 microM for arsenite and 132 microM for arsenate.;In Chapter 3, a mitochondrial paper-based biosensor was fabricated. Coupled mitochondria were isolated from bovine heart and demonstrated an amperometrical detection limit of 20 nM for malathion, a common pesticide. The inhibition mechanism by malathion to mitochondrial metabolism was studied electrochemically and was determined to be uncoupling rather than inhibition.;In Chapter 4, the inhibition mechanism of mitochondria by rotenone, carboxin, and antimycin was studied. It was also discovered that the synergy between riboflavin derivatives and ubiquinone can be altered by using different solvents during the electrode fabrication process. A further study indicated that lipid membrane is capable of altering the reaction dominance between ubiquinone and riboflavin derivatives. Finally, it was discovered that mitochondria release riboflavin derivatives under inhibition and it is believed that this alteration of the micro-environment was the cause of the change in mitochondrial electrochemistry when mitochondria were inhibited.