Preparation And Electrochemical Characteristics Of Novel Tyrosinase Biosensors

A novel tyrosinase carbon paste biosensor (T-CPE-IL) was constructed with paraffin oil and ionic liquid, 1-butyl-3-methyl-imidazolium hexafluorophosphate ([BMIM]PF6), as binder to immobilize tyrosinase. Then, a sodium alginate modified tyrosinase carbon paste biosensor(SA-T-CPE-IL) based on T-CPE-IL was also developed. The effect of [BMIM]PF6 on the electrochemical behaviors of tyrosinase carbon paste electrode (T-CPE) was studied with cyclic voltammograms, chronocoulometry and alternative current impedance spectroscopy. Finally, T-CPE-IL has been successfully used for determining the trace amounts of phenol in practical environmental water samples.The main points of the dissertation were as follow:1. Preparation of T-CPE-IL. Under the optimized parathion conditions, [BMIM]PF6/binder =85%(w/w) and enzyme/carbon paste =7%(w/w), T-CPE-IL could present higher sensitivity,better stabilization and faster response for catechol than that of T-CPE. In pH=6.5 sodium phosphate buffer solution, the electrochemical behaviors of T-CPE-IL were studied with cyclic voltammograms and chronocoulometry. Experimental results showed that the reductive currents were increased with the scan rates, Ipc = 0.0016v + 0.1955,R2 = 0.9910,which indicated that the redox process was surface-controlled. Moreover, the response time of the electrod was less than 10s. In addition, the concentrations of catechol in the range of 20μM～200μM were linear with the increase of the reductive currents of T-CPE-IL, while the detection limits of catechol was of 3μM.2. Effect of [BMIM]PF6 on the electrochemical behaviors of T-CPE. FT-IR spectra,Chronoamperometry,Cyclic voltammograms and alternative current impedance spectroscopy were used to study the effect of [BMIM]PF6 on the secondary structure of tyrosinase,the adsorption performance of catechol and the standard rate constant of electron transfer of T-CPE-IL. Experimental results showed that T-CPE-IL could influence the secondary structure of tyrosinase, inprove the surface characteristics of T-CPE and increase the adsorption capacity of catechol on the surface of the electrode. The adsorption capacity of catechol on the surface of the electrode was increased from 2.08 mc to 5.67mc and the standard rate constant of electron transfer was increase more than 2 magnitudes. Moreover, the process of the electrode reaction was changed from electron transfer control to diffusion control.3. Manufacture of sodium alginate modified tyrosinase carbon paste biosensor (SA-T-CPE-IL). The electrode components and some analytical characteristics of SA-T-CPE-IL were studied. The effects of sodium alginate on the electron transfer were studied with cyclic voltammograms and impedance spectroscopy. Experimental results showed that the response of catechol on SA-T-CPE-IL was more reversible than that of T-CPE-IL. However, the standard rate constant of electron transfer was decreased and the process of electrode reaction changed from diffusion control to electron transfer control due to the fact that the unconductivity of sodium alginate. Moreover, the concentrations of catechol in the range of 80μM～400Μm were linear with the increase of the reductive currents of T-CPE-IL.4. Comparation of the analytical characteristics of T-CPE-IL and SA-T-CPE-IL. The tolerated amounts of common co-existing substances on the determination of catechol in practical environmental water samples were determined. In addition, T-CPE-IL could be used to determine the trace amounts of catechol in practical environmental water samples with satisfactory results.