Electrochemical growth of polypyrrole based nanostructures for development of enzymatic amperometric glucose biosensors

Abstract

Glucose is a key analyte in the elds ranging from food industry to biomedical analysis. It plays a very important role in human metabolism. Hence, monitoring and control of glucose concentration in human body is of great signi cance. Biosensors have emerged as one of the most reliable tools among the existing conventional techniques for assay of glucose. The research in the eld of biosensors has been accelerated in the past few decades for various applications such as glucose monitoring, drug discovery, drug anal- ysis and food analysis etc. Of these, more than 85% of biosensor market is captured by glucose biosensors due to tremendous increase in the number of diabetic patients worldwide. Owing to extraordinarily superior speci city and sensitivity of enzymes to- wards speci c substrate molecules accelerated research in the eld of enzymatic glucose biosensors has been encouraged among researchers. Glucose detection has been re- portedly performed using various techniques viz. chromatography, chemiluminescence, electrochemistry etc. Among these, electrochemical methods, especially amperometric detection has been proven to be one of the most simplistic and most powerful method- ologies. Conducting polymer nanostructures have been proved to be materials of choice due to their excellent electronic properties and biocompatibility. In this work, we have fabricated a variety of amperometric enzymatic glucose biosen- sors based on growth of polypyrrole (PPy) nanostructures using two di erent ap- proaches: (1) template based appraoch, (2) template free approach. In both the ap- proaches PPy nanostructures were grown using potentiostatic electropolymerization employing LiClO4 as supporting electrolyte and retained the very essential features of green chemistry. In the rst approach, PPy nanotube arrays were grown on alumina membranes (anodiscTMs) with two pore diameters: 100 and 200 nm. Platinum coatedAnodiscTM was used as working electrode for electrodeposition in the aqueous solution containing pyrrole monomer and supporting electrolyte. The electrodes so obtained were used as supporting matrix for glucose oxidase (GOx) immobilization. By varying polymerization time during the potentiostatic electropolymerization, the size/diameter of the PPy nanotubes were controlled, leading to changes in the subsequent enzyme immobilization. As improvements in the sensitivity of biosensors have always been of paramount interest, polymerization time was optimized for the highest sensitivity, in each case. For this purpose, immobilization of GOx was rst performed using physi- cal adsorption and biosensing response was examined amperometrically for increasing concentrations of glucose. Also, quanti cation of immobilized enzymes was performed using uorescence spectroscopy. In order to further improve the sensing performance and storage stability of the biosensor fabricated for optimum polymerization duration, GOx immobilization was carried out using cross linking with glutaraldehyde and bovine Serum Albumin. Manifold enhancements in the sensitivity and shelf life were observed in the fabricated electrodes. The biosensors also showed extended linear range of oper- ation and satisfactory low detection limit. Template free approach for electropolymerization resulted in to PPy nano ber net- work. The main aim of this research was to develop a template free method involving minimum number of process steps and mild chemicals, for obtaining CP nanostructures with controllable size and morphologies. As intended, a simple, economic, template free, single step potentiostatic electropolymerization method has been developed for the growth of porous PPy nano ber network. PPy was grown on gold coated glass substrates. The critical electrosynthesis parameters viz. monomer and supporting electrolyte concentrations and polymerization potential were optimized for the highest sensitivity of the fabricated biosensors. This is the rst report, which demonstrates the application of the PPy nano bers fabricated under the chosen synthesis condi- tions, towards enzymatic glucose biosensor. The observed results indicate that low monomer concentration and moderately low polymerization potential are highly suit- able for growth of PPy nano ber network with controllable ber diameters having high aspect ratio, resulting in high sensitivity for biosensor application.