Construction Of Protic Ionic Liquid Modified Electrode And Its Enhanced Action In Direct Electron Transfer Of GOD

Ionic liquids (ILs) are molten salts with the melting point close to or below room temperature. ILs have many unique chemical and physical properties, such as high chemical and thermal stability, good ionic conductivity, negligible vapor pressure and wider electrochemical windows. Recently, ILs are attracting intensive interest in the electrochemistry and electroanalysis field due to their high electrochemical stability, ionic conductivity. Here, a PIL (protic ionic liquid), N-ethylimidazolium trifluoromethane sulfonate ([EIm][TfO]), was chosen as an effective modifier to fabricate a novel GOD(glucose oxidase) biosensor. The direct electrochemistry and electrocatalytic behaviour of GOD on the biosensor was investigated. The Further improvement of the experiment was considering the benefits of CHIT (chitosan), ILs, CNTs (multi-wall carbon nanotubes), we have integrated them in a biosensor fabrication to exploit their synergistic contributions on the improvement of sensor characteristics.The main work and research results are as follows:In the first part of this paper, the best optimization kind of IL was chosen and the conditions of CILE formation were studied. This paper, we chose carbon paste electrode as foundation electrode and PIL,paraffin as binder. We chose four kinds of ILs (N-ethylimidazolium trifluoromethane-sulfonate ([EIm][TfO]), 1-butyl-3-methyl- imidazolium tetra?uoroborate (BMIMBF4), N-butylpyridinium tetra?uoroborate (BPBF4), Alkyl quaternary ammonium tetra?uoroborate) for compared. The results proved [EIm][TfO] can improve the reversibility and the stability of CILE, the electrode had the lowest background current and the highest conductivity, so [EIm][TfO] was chosen for the subsequent work. The conditions of CILE formation were studied and optimized, the effect of PIL content upon cyclic voltammograms (CVs) in a 5 mM potassium ferricyanide solution showed that the 2/13 ratio of PIL and paraffin was the most optimization ratio.In the second part of this paper, we modified GOD on the CILE by Nafion film to obtioned Nafion/GOD/CILE. The SEM image showed the immobilized GOD molecules was highly dispersed and showed a uniform snowflakes structure on the surface of CILE. Cyclic voltammetry (CV) was used to investigate the electrochemical behavior of the modified electrode and bioelectrocatalytic activity of enzymes, a pair of well-defined and nearly symmetric redox peaks for the direct electron transfer of GOD could be observed. The redox peak potentials were -0.437 and -0.515 V at 100 mV s^-1, its formal potential (E⁰′) was around -0.476 V. The cyclic voltammograms of Nafion/GOD/CILE at various scan rates were investigated, the rate constant ks was ca. 3.98s?1, the average surface coverage (Γ*) of GOD was calculated to be 3.077×10^-10 mol?cm^-2 from the slope of the Ip-νcurve. Cyclic voltammograms of Nafion/GOD/CILE showed a strong dependence on solution pH, E⁰′had a linear relationship with pH (the linear regression equation is E⁰′= -0.174-0.0456 pH, R = 0.998), the slope was nearly close to the theoretical value (-58.6mV/pH). It proved that two protons and two electrons participate in the electrochemical reaction of the GOD immobilized on CILE. The further experiments proved that the bisensor has a good corresponding signal change with glucose concentrations. The linear response was in the range 0.005-2.8 mM, the sensitivity and the apparent Michaelis-Menten constant (Km) were estimated to be 14.96μA mM^-1 and 1.53 mM. All experimental results suggested that the use of protic IL [EIm][TfO] can retain the bioactivity of GOD and enhance the sensitivity and the stability of the biosensor. The proper reason was that PIL can provide a special micro-envieonment for GOD. Nafion/GOD/CILE was used to detection the glucose concentration in fresh human serum samples, the results were in good agreement with hospital method. All the results demonstrated the reliability of the fabricated PIL-based biosensor. The good properties of the modified electrode implied that PILs could be applied to provide a promising strategy for the development of biosensors.In the third part of this paper, we further modified CHIT,CNTs on the CILE to obtioned GOD/CNTs-CHIT/CILE. The electrochemical behavior and the electrocatalytic activity of GOD were investigated liked the second part of this paper. Cyclic voltammograms showed the redox peaks of GOD/CNTs-CHIT/CILE is 3 times more than Nafion/GOD/CILE. The rate constant ks was ca. 5.836s^-1 and the average surface coverage (Γ*) of GOD was calculated to be 6.887×10^-10 mol·cm^-2, both were 2 times more than Nafion/GOD/CILE. Current-time curves showed the biosensor has a rapid and sensitive reponse to changes of glucose concentration, the linear response was in the range 0.04-2.8 mM with the sensitivity of 35.5μA mM^-1cm^-2. The apparent Michaelis-Menten constant (Km) was 1.208 mM, which was smaller than the number of Nafion/GOD/CILE. All experimental results suggested that the modified GOD has high bioactivity due to the synergistic influence of CHIT, ILs and CNTs. The method presents here can be easily extended to immobilized and obtain the direct electrochemistry of other enzymes or proteins.