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Role of Specific Enzymes in Biosensor for Tea Analysis

Sujith Kumar, P. V. (2010) Role of Specific Enzymes in Biosensor for Tea Analysis. PhD thesis, University of Mysore.

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Abstract

Polyphenol oxidase (PPO) from tea leaves, potato, brinjal, apple and mushroom were assessed for their selectivity towards PP (catechin-a specific PP present in tea leaves). PPO from tea leaves was purified to homogeneity with three isozymes. PPO from other sources also partially purified and compared with standard tyrosinase and laccase. Among all these enzymes tyrosinase found to have highest substrate specificity towards catechin. This was further confirmed by enzyme kinetics studies. In the immobilized condition PPO from tea leaves and tyrosinase were responded to catechin. For black tea sample analysis by tea biosensor, only tyrosinase was given measurable signals, but for green tea sample analysis both tyrosinase and PPO were found to be good. A detector system comprising of two electrodes was fabricated in-house and used for the analysis of PP using enzyme based biosensor. It is programmed to detect at various concentrations of PP with good sensitivity. Immobilized enzyme was stabilized by various PBSAs and lysozyme observed with greater stability and enhanced the number of analysis. Other physico-chemical parameters were optimized. With this conditioning biosensor could analyze about 100 samples by using single immobilized enzyme membrane. During PP analysis it was observed that immobilized enzyme membrane was inactivated due to fouling. This fouling was attributed to protein- PP interaction. Protein-PP interactions were studied using various amino acids. Amino acid residues such as lysine, proline and serine observed with maximum affinity with PP. 10 mM concentration of PP found to be the optimum substrate concentration to minimize the fouling. Dissociation studies were carried out with various dissociating agents. Glycerol found to be better polyol to prevent the complete binding of PP with proteins. Developed tea biosensor exhibited good robustness in a number of in lab and field trials. Commercial black tea samples and black tea grades were analyzed for their PP content to evaluate its quality. ii Biosensor system was validated by using various conventional analytical techniques. Biosensor system analysis also validated by tea taster’s score for various grades of tea. In all these studies biosensor has shown a good positive correlation with other conventional analytical techniques. Tea biosensor developed as an outcome of this research work could be a potential tool to analyze tea quality based on PP content, hence major quality attributes of tea such as colour and astringency more or less derived from PP present in tea. This methodology is rapid, simple, user friendly and cost effective. It also helps in quality monitoring at different stages of processing (inline monitoring) and could be adapted for the on field evaluation of quality of tea.

Item Type: Thesis (PhD)
Uncontrolled Keywords: Polyphenol oxidase, tea leaves, biosensor
Subjects: 600 Technology > 05 Chemical engineering > 01 Biotechnology and Bioengineering
600 Technology > 07 Beverage Technology > 08 Tea
600 Technology > 08 Food technology > 16 Nutritive value > 05 Enzymes
Divisions: Fermentation Technology and Bioengineering
Depositing User: Food Sci. & Technol. Information Services
Date Deposited: 15 May 2012 10:59
Last Modified: 15 May 2012 10:59
URI: http://ir.cftri.com/id/eprint/10763

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