Fabrication Of Laccase Electrode Based On Functionalized Conducting Polymers And The Direct Electrochemistry

Laccase is a multi-active-center glycoprotein.It catalyzes the oxidation of phenols or aromatic amine substrates with the concomitant reduction of molecular oxygen into water.This catalytic feature gives its potential value not only in the field of environment protection but also in the fields of biofuel cell and biosensor.In order to obtain high performance laccase electrode,it is necessary to shorten the distance between the laccase active site and the electrode surface,and reduce the dependence of the direct electron transfer process on laccase orientation.In order to achieve the above objectives,a new immobilization strategy must be developed.The chemical affinity strategy and protein recombination strategy have been reported for laccase immobilization.Compared to the biochemicalor molecular biology modification of laccase,it is also a good way to construct a nanostructured carrier which is suitable for the efficient heterogeneous electron transfer of laccase.The use of conducting polymers to prepare laccase cathodes has been tried.The major problem is the low laccase loading.Laccase is a glycoprotein,its outer glycosyl group could be used as immobilization sites.Laccase can be immobilized on the electrode surface based on the reaction of its glycosyl group with the boronic acid group of the phenylboronic acid type functionalized conducting polymers.A conducting polymers-based sandwich-type laccase electrode could reduce the dependence of the direct electron transfer process on laccase orientation due toits cage effect,and further improve the electron transfer efficiency of laccase,and therefore the performance of the laccase electrode.Based on the above background,I carried out two works,which are summarized and presented as follows:1.Electrochemical synthesis of poly(3-aminophenylboronic acid)in ethylene glycol without exogenous protonsA non-aqueous solution of tetra-n-butylammonium fluoride(TBAF)in ethylene glycol has been tried for the first time as a supporting electrolyte for the electropolymerization of 3-aminophenylboronic acid(APBA).Unlike the traditional acidic aqueous solution,the present medium needs no exogenous protons;moreover,the presence of CF3COOH is found to be unfavorable for the polymerization.The protons are in situ generated by the reaction between the boronic acid group on APBA and 1,2-dihydroxyl on ethylene glycol.So ethylene glycol serves as not only solvents but also proton sources.As a part of the supporting electrolyte,F-is found to be involved in the electrochemical synthesis of poly(3-aminophenylboronic acid)(PAPBA),but it is not indispensable.Studies on the electropolymerization process indicates that the size of the ions in the electrolyte affects the rate of the doping/dedoping process.The smaller the cation,the easier the doping/dedoping process,and the better the stability of the grown film.As demonstrated by Fourier transform infrared spectra,UV-Vis spectra,and scanning electron microscopy,the obtained PAPBA is a cross-linked nanoporous polymer membrane that has good adherence to the glassy carbon electrode.2.Construction and Characterization of sandwich-type laccase electrode based on functionalized conducting polymersLaccase(Lac)is a multi-active-center glycoprotein.The heterogeneous electron transfer efficiency of laccase depends on the distance between the substrate binding site and the electrode surface.Laccase encapsulated by conducting polymer should be favorable for its direct electron transfer process,thus improving the electrocatalytic efficiency of the laccase electrode.Here,a sandwich-type laccase electrode based on functionalized conducting polymers was developed.Firstly,porous poly(3-aminophenylboronic acid)(PAPBA)conducting film was deposited on a glassy carbon electrode(GCE)by cyclic voltammetry,and on which laccase was thenimmobilized via the specific binding of the boronic acid group from PAPBA and the glycosyl group on laccase.In addition to higher laccase loading,the present Lac/PAPBA/GCE electrode is more robust than that prepared by physical adsorption of laccase on conducting polymer.For laccase to be wired better,a poly(3,4-ethylenedioxythiophene)(PEDOT)conducting film with polystyrenesulfonate anion(PSS-)as the dopant was covered on the Lac/PAPBA/GCE electrode by potentiostatic electrodeposition.The electrical activity as well as the biological activity of the laccase encapsulated by the dual layer polymer depend on the electrodeposition conditions.Under the same conditions,the sandwich-type laccase electrode produces less H2O2 than the laccase electrode without the second layer of PEDOT conducting polymer;i,e.,the introduction of the second PSS doped PEDOT film can reduce the dependence of the DET process on laccase orientation and improve the efficiency of the electroreduction of O2.In addition,the sandwich architecture can reduce the requirement for the stability of the cyclic boronic acid ester and improve the biological activity of the laccase and the polymer conductivity,thereby enhancing the bioelectrocatalytic performance of the laccase’electrode for oxygen reduction.