Please use this identifier to cite or link to this item: http://hdl.handle.net/10266/5993
Title: Development of Some Biosensors and Their Analytical Applications
Authors: Pabbi, Manisha
Supervisor: Mittal, Susheel
Keywords: Whole cell biosensors,;ZnO nano particles;pesticide;acephate;chlorpyrifos
Issue Date: 7-Aug-2020
Abstract: The present work provides an insight into the concept, development, and application of the whole-cell biosensors for organophosphorus pesticide determination. For the fabrication of biosensor, Chlorella sp. algal cells having phosphatase activity were chosen. The reason for choosing algal cells as a sensor-modifying agent was their stability in quasi-physiological conditions, easy to handle, economical, high productivity and have a longer life span compared to bacterial or other whole cells. It is necessity for a biosensor that biological recognition element and transducer should work as a single unit, thus, immobilization becomes an important step for biosensor development. Our work has primarily focused on the fabrication of electrochemical biosensors. The algal cells with anticipated phosphatase enzyme were immobilized on the electrode surface using appropriate techniques. The immobilization technique solely depends upon the type of electrode used for preparation of biosensor. The thesis has been divided into two sections- (i) Inhibition based biosensors, (ii) Catalytic biosensors. Inhibition based biosensor are those in which pesticide concentration is determined by measuring the extent of inhibition of the enzyme activity. To increase sensitivity, operational stability, and to attain low detection limit, ZnO nanostructures have been introduced in the design of biosensor. Two types of biosensors have been developed under this section: (i) An electrochemical biosensor for the determination of acephate pesticide was developed based on algal modified silica coated ZnO quantum dots (QDs). The surface active alkaline phosphatase enzyme on the Chlorella sp. algae cell wall dephosphorylates substrate p-nitrophenyl phosphate (pNPP) and release p-nitrophenol (pNP), which being electroactive gets oxidized on the modified working electrode to generate current equivalent to the amount of pNP released from the substrate. The presence of acephate would inhibit formation of pNP and hence decrease in current, which indirectly corresponds to the concentration of the pesticide present. The biosensor responds linearly in the concentration range of 10−11 M to 10−3 M for acephate and showed QDs strongly influenced the biosensor performance as they increased sensitivity and improved the detection limit. (ii) An algal based biosensor modified with flower shaped ZnO nanoparticles for the determination of chlorpyrifos in an aqueous medium was developed. Enzyme would dephosphorylate the phosphate monoester of substrate 2-phospho-L-ascorbic acid to release L-ascorbic acid, which was monitored using voltammetric and ISFET techniques. The presence of pesticide would inhibit AP-algae enzyme activity and hence the current magnitude decreases. The decrease in current is quantitative in nature and used for determination of concentration of the pesticide. Flower shape ZnO nanoparticles not only provide surface for adsorption of AP-algae for better immobilization but also increases electron-transfer kinetics and sensitivity of the biosensor. The voltammetric and ISFET methods could measure chlorpyrifos in the linear concentration range from 10-9 M to 10-1 M and 10-10 M to 10-3 M, respectively, with nil interference from triazophos, malathion, acephate and some metal ions. Application aspects of the proposed biosensor were demonstrated using extracted soil samples and showed excellent reproducibility and reliability. Catalytic biosensors use pesticide as a substrate. Enzyme has the capability to degrade the pesticide by hydrolyzing a specific bond. Under this section, a screen-printed electrode based algal biosensor was developed for the detection of chlorpyrifos. Alkaline phosphatase acts upon chlorpyrifos and degrade it into 3,5,6-trichloro-2-pyridinol, which is monitored through voltammetry and chronoamperomerty. It was observed that the voltammetric method could determine chlorpyrifos up to 10-7 M whereas chronoamperometric method could detect upto 10-10 M concentration. Spectrophotometric and HPLC studies were also performed to validate the results obtained from proposed biosensor.
URI: http://hdl.handle.net/10266/5993
Appears in Collections:Doctoral Theses@SCBC

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