CONTENT OVERVIEW Chapter 1 Overview Chapter 2 Peroxide Activation using Copper-Induced Fenton-Like Catalysis Chapter 3 Effects of Matrix and Ligand on the Activation of Hydrogen Peroxide with Copper(II)-Amino Acid Complexes Chapter 4 Characteristics of Peroxidase-Entrapped Silica Modified with Covalently Bound Sugars for Enhancing Biocompatibility Chapter 5 Controlling Structural Characteristics and Biological Activity of Peroxidase- Entrapped Bio- glasses via PEG Polymer Doping Chapter 6 Future Outlook: Measurement of Peroxide and Antioxidant Activity Using HRP-Entrapped Sensor and Monolithic Silica Capillary Column combined with UV Detection CONTENTS Abstract (Chinese) I Abstract (English) II Content Overview III Chapter 1 Overview 001 Chapter 2 Peroxide Activation Using Copper-Induced Fenton-Like Catalysis 006 2.1 Introduction and Research Goal 006 2.2 Background 007 2.2.1 Fenton reaction/ Fenton-like reaction 007 2.2.2 The nature of copper ions and copper complexes 008 2.2.3 Peroxide-activation pathway and its application in engineering 009 2.3 Experimental 012 2.3.1 Reagents 012 2.3.2 Instruments 012 2.3.3 Procedures 013 2.4 Results and Discussion 015 2.4.1 Kinetics of substrate oxidation 015 2.4.2 pH effect 023 2.4.3 Effect of [copper: amino acid] molar ratio 026 2.4.4 Analysis of copper-complex activity vs. copper speciation 029 2.4.5 Side-chain effect on the copper-complex activity 031 2.5 Conclusions and Future Outlook 032 2.6 References 035 Chapter 3 Effects of Matrix and Ligand on the Activation of Hydrogen Peroxide with Copper(II)-Amino Acid Complexes 041 3.1 Introduction and Research Goal 041 3.2 Background 042 3.2.1 The importance of copper in radical-generating system 042 3.2.2 The role of amino-acid chelator on the activity of copper(II) complexes 043 3.2.3 The role of antioxidant on the activity of copper(II) complexes 046 3.2.4 The investigation of reactionpathways on Cu(II)/organic acid/H2O2 system 047 3.3 Experimental 048 3.3.1 Reagents 048 3.3.2 Instruments 048 3.3.3 Preparation and procedures for kinetic study 049 3.4 Results and Discussion 051 3.4.1 Effects of buffer concentration and ion strength 051 3.4.2 Effect of electrolytes and their concentrations 055 3.4.3 Effect of amino acid chelators 059 3.4.4 Competitive reaction between antioxidants and dyes 061 3.5 Conclusions and Future Outlook 066 3.6 References 068 Chapter 4 Characteristics of Peroxidase-Entrapped Silica Modified with Covalently Bound Sugar for enhancing Biocompatibility 073 4.1 Introduction and Research Goal 073 4.2 Background 074 4.2.1 Characteristics and function of HRP 074 4.2.2 Development and characteristics of silica matrix using sol-gel technique 076 4.2.3 Immobilization of enzymes in silica matrix using one-step sol-gel technique 078 4.2.4 Developed methods using HRP as the catalytic medium 081 4.3 Experimental 083 4.3.1 Materials and reagents 083 4.3.2 Preparation of DGS 083 4.3.3 Preparation and procedures 083 4.3.4 HRP activity assay 084 4.4 Results and Discussion 085 4.4.1 Interference of additives for HRP activity assay in solution 085 4.4.2 Kinetics of HRP activity assay in solution 088 4.4.3 Effect of immobilization on catalytic activity of encapsulated HRP 091 4.4.4 Effect of aging condition on encapsulated HRP performance 093 4.4.5 Optimization of composition of silica monoliths 096 4.4.6 Sugar-modification on biocompatibility of silica (DGS vs. SS) 099 4.4.7 Effect of timing of GLS addition 104 4.5 Conclusions and Future Outlook 110 4.6 References 111 Chapter 5 Controlling Structural characteristics and Biological Activity of Peroxidase-Entrapped Bio-Glasses via PEG-Polymer Doping 118 5.1 Introduction and Research Goal 118 5.2 Background 119 5.2.1 Functions of PEG organic modifiers on enzyme activity 119 5.2.2 Function of organic modifiers on silica morphology 120 5.2.3 Application of PEG polymer-modified silica medium 121 5.2.4 PEG-modified medium as biocompatible materials 122 5.3 Experimental 124 5.3.1 Reagents 124 5.3.2 Experimental 124 5.3.3 Characteristics evaluation of silica morphology 124 5.4 Results and Discussion 126 5.4.1 Morphology investigation on PEG polymer-doped silica 126 5.4.2 Effect of PEG additives on leaching of HRP from silica gels 132 5.4.3 Encapsulated HRP activity in different silica monoliths 134 5.5 Conclusions and Future Outlook 137 5.6 References 138 Chapter 6 Future Outlook: Measurement of Peroxide Using HRP-Entrapped Silica Medium and Monolithic Silica Capillary Column Combined with UV Detection 143 6.1 Introduction and Research Goal 143 6.2 Background 144 6.2.1 Silica-derived monolith as bioreactor and liquid chromatography column 144 6.2.2 Applications of HRP-encapsulated silica matrix in chemical assay 145 6.2.2 Functions of radical scavengers/antioxidants and its measurement 146 6.3 Experimental 149 6.3.1 Reagents and Instruments 149 6.3.2 Procedures for column making 149 6.3.3 Analytical procedures 151 6.4 Results and Discussion 153 6.4.1 Kinetic parameters of entrapped HRP in different silica materials prepared in 96-well microtiter plates 153 6.4.2 Measurement of peroxide using array-type optical sensor 193 6.4.3 Effect of electrostatics on the entrapping efficiency and stability of HRP 159 6.4.4 Stability and reusability of synthesized HRP reactor 163 6.4.5 Stability of ABTS radicals formed in the flow-through HRP reactor 165 6.5 Conclusions and Future Outlook 170 6.6 References 172