Abstract
The accurate and rapid determination of hydrogen peroxide (H2O2) is of great importance in many applications such as clinical diagnosis, bioanalysis, and food safety. Moreover, H2O2 is a side product of specific enzymatic reactions. For instance, glucose could be oxidized to produce gluconic acid and H2O2 in the presence of glucose oxidase (GOx). Up until now, there have already been many methods to detect H2O2, including spectrophotometry, electroanalysis, and fluorometry. However, these techniques require expensive or sophisticated instruments. Therefore, rapid and easy ways of H2O2 sensing are necessary. In this thesis, we have developed two systems to detect H2O2 without any complicated equipment. Both of them are simple, low cost, and disposable. Most importantly, we could distinguish the color change by the naked eye. By these advantages, the sensor could be applied to develop point-of-care (POC) and clinical diagnosis platforms. The intrinsic catalytic ability of palladium in many specific reactions has been confirmed. Herein, our first system utilizes the enzymatic approach by using hollow silver/palladium nanostructures (Ag/Pd NSs) to replace commercial enzyme, Horseradish peroxidase (HRP), to catalyze the oxidization of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) by H2O2. By calculation of many kinetic constants from enzyme kinetics, we compared the catalytic activity of Ag/Pd NSs with HRP. Proved that the Ag/Pd NSs has high catalytic activity and similar results to HRP. We also did the catalytic reaction in different reaction conditions such as temperature, pH value and reaction time. The most suitable reaction conditions were under room temperature, acidic condition (pH=4.6) and 10 min in solution. Then, we have successfully immobilized a nanoparticle array onto the commercial polymer substrate, polyethylene terephthalate (PET), via a wet-chemical method. By mixing H2O2 with ABTS in an acetic buffer (pH=4.6) with the Ag/Pd NSs-coated PET sensing system, we could observe the increase in darkness of green color upon increasing H2O2 concentration by the naked eye. The high catalytic activity of Ag/Pd NSs might be attributed to the hollow structure (large surface area) and the intrinsic catalytic ability of palladium. The linear relationship was from 5 mM to 500 mM in solution-based test and 2 mM to 10 mM in PET-based detection system. In another system, we immobilized hollow silver/gold nanostructures (Ag/Au NSs) onto PET substrates, similar to the first system. The detection process could be performed by directly dropping the H2O2 solution onto the Ag/Au NSs-coated PET sensing system, without any organic dyes. Because of the strong oxidizing ability of H2O2, the silver atoms in the Ag/Au NSs would be oxidized to silver ions and the Ag/Au NSs would transform to a more porous structure. Owing to the changes in structure and composition, the localized surface plasmon resonance (LSPR) peak of Ag/Au NSs would continuously shift to longer wavelength (red shift) and different colors would be obtained at various H2O2 concentrations, visible by the naked eye. We could successful detect the H2O2 under room temperature and neutral condition (pH 7.0) in 30 min. We obtained narrow linear range but low detection limit, 5 µM to 50 µM and 4.3 µM respectively. Finally, the Ag/Au NSs-based platforms could be utilized for clinical diagnosis, such as glucose, cholesterol or uric acid sensing. In this study, we have successfully combined the cholesterol oxidase to develop cholesterol sensing. The linear range was from 10 µM to 50 µM. The detection limit was low as 6.0 µM. An overview of the two efficient sensing platforms, the second one shows lower detection limit but narrow linear range than first one. In the future, the applications of the sensing systems could extend to other fields that relate to H2O2.