Studys On Biofunctionalization Of Electrode Interface Via Click Chemistry

Click chemistry is proposed by Sharpless(winner of the nobel prize in chemistry 2001) and his coworkers in 2001.Click chemistry enables a modular approach to generate novel moleculars utilizing a collection of simple and reliable chemical reactions.It creates a new aspect of combination chemistry which can generate novel products, fastly,effectively and stereoselectively in high yields.Click chemistry's applications are increasingly found in aspects of drug discovery, polymer science and bioconjugate chemistry.But it is limitedly used to functionalize the solid phase of chromatography in analytical chemistry.Thus,the further development of the application of click chemistry in analytical chemistry is very important.It is a new kind of research field to apply click chemistry to biofunctionalize the electrode surfaces and to develop a novel strategy that can attach moleculars with controlled surface density.We try our best to combine click chemistry,life science with electroanalytical chemistry firmly.The paper concludes following aspects:Chapter one:PerfaceBasic concept,characteristic and reaction types of click chemistry are introduced.The applications of click chemistry in drug discovery,polymer science,bioconjugate chemistry and functionalization of solid surface are summarized.Chapter two:Covalent grafting nitrophenyl group on Au surface via click reaction:Assembling process and electrochemical behaviorsIn this paper,we described a simple and facile method for the covalent functionalization of Au surface with nitrophenyl group via stepwise strategy. Structurally well-defined azide-terminated organic selfassembled monolayers(SAMs) were formed on Au surface from a commercially available mixture solution of azidoundecanethiol and dilute thiol.Subsequent,derivatization of the azide-terminated monolayers was conducted in aqueous environments with ethynyl nitrobenzene via a selective,reliable,robust click reaction.By this way,the nitrophenyl group was covalently and quantitatively grafted on Au surface,which was confirmed by Raman Spectrometry and electrochemical methods.These results demonstrated the efficiency of using click chemistry in assembling covalently linked nanostructures.Nitro groups were transformed to amines via electro-reduction and subsequently crosslink with biomacromolecules for solid surface biofunctionalization. The direct electron transfer of hemoglobin(Hb) was observed.And the Hb modified electrode exhibited a good electrocatalytic performance for H₂O₂.Chapter three:Immobilization of protein via click reaction and its electrochemistryIn this paper,we have prepared mixed self-assembled monolayers(SAMs) on gold electrode from azido-terminated alkane thiols and alkane thiols,then,hemoglobin(Hb) was immobilized on the electrode through click reaction and carbodiimide reaction. Reflectance absorption infrared(RAIR) spectroscopy,Electrochemical impedance spectroscopy(EIS),and Cyclic voltammetry(CV) were applied to characterize the modified electrode.Cyclic voltammetry of Hb modified electrode showed a pair of well-defined and quasi-reversible peaks for Hb Fe(â…¢)/Fe(â…¡) redox couple at about -0.210V(V.S.SCE) in 0.1 mol/L pH 5.0 buffers.The number of electrons transferred per protein molecule was 0.75,and the electron transfer rate constant(k_s) was estimated to be 0.779s^-1.When the various mole fraction of azido-terminated alkanethiol in the deposition solution(x_soln) is 0.5,the surface coverage(Γ) of the immobilized Hb reached a maximal value(8.3×10^-12 mol·cm^-2),indicating the formation of a monolayer.The modified electrode displayed a good electrocatalytic response to the reduction of O₂ and H₂O₂,Its linear range were 3.5 mmol/L to 34.8 mmol/L and 1.0μmol/L to 0.375 mmol/L,respectively.The apparent Michaelis-Menten constant was calculated to be 0.107 mmol/L,suggesting that Hb molecules immobilized on the modified gold electrode retained their bioactivity.Chapter four:Preparation of tyrosinase modified electrode via click chemistry and its application in phenol detectionCombined SAM and click reaction,a nitrophenyl modified Au surface was produced. The nitrophenyl group terminated SAMs were conversed to aminophenyl group terminated SAMs by electro-reduction.And then the tyrosinase was covalently immobilized onto the Au electrode via carbodiimide reaction.Cyclic voltammetry (CV),Electrochemical impedance spectroscopy(EIS),and Reflectance absorption infrared(RAIR) spectroscopy were used to characterize the modified electrode.The tyrosinase modified electrode displayed a good electrocatalytic response to the reduction of phenol,pyrocatechol and m-Cresol,ks linear range was 0.2μmol/L to 6.0μmol/L,0.2μmol/L to 73.1μmol/L,and 0.2μmol/L to 53.0μmol/L,respectively. The apparent Michaelis-Menten constant was calculated to be 0.032,0.038 and 0.055 mmol/L,respectively.The tyrosinase modified electrode was used for determination of phenol in the tap water using the standard method.