Studies On The Electrochemical Behaviors Of Cytochrome C On2-mercaptopyrimidine Derivatives Modified Gold Electrode

The preparation and promotion of2-mercaptopyrimidine derivatives and2-mercaptonicotinic acid modified gold electrodes on the direct electrochemical reaction of cytochrome cwere investigated by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS),scanning electron microscopy (SEM), UV-visible spectroscopy (UV-vis) and amperometry.The effects of modification and electrochemical conditions, molecular structure of2-mercapto-pyrimidine derivatives were discussed.1. The electrochemical properties of bare gold electrode and2-mercaptopyrimidinederivative modified electrodes were compared in K3[Fe(CN)6] solution. It was found that thereversibility of the electrochemical reaction of K3[Fe(CN)6] was decreased on the2-mercaptopyrimidine derivative modified electrodes. The peak potential difference wasincreased, and the peak current was decreased.Relative uniform self-assembled film of2-mercaptopyrimidine derivatives formed on thesurface of modified gold electrode was characterized by scanning electron microscopy (SEM).Uniformly dispersed cytochrome c absorbed on the surface of4-hydroxy-2-mercapto-5-methylpyrimidine or2-mercaptonicotinic acid modified electrode were observed, which hadremarkable catalytic effect on the electrochemical reduction of H2O2, and can be used to thepreparation of electrochemical biosensor for the determination of H2O2.2. A series of2-mercaptopyrimidine derivatives and2-mercaptonicotinic acid was found tohave good promotion effect on the direct electrochemical reaction of cytochrome c. Theoptimum promotion effect of2-mercaptopyrimidine,4-amino-2-mercaptopyrimidine,4-hydroxy-2-mercapto-5-methylpyrimidine or4-amino-6-hydroxy-2-mercaptopyrimidine modifi-ed gold electrode on the direct electrochemical reaction of cytochrome c was obtained byimmersing the gold electrode in2.0mmol/L2-mercaptopyrimidine,4-amino-2-mercapto-pyrimidine,4-hydroxy-2-mercapto-5-methylpyrimidine or4-amino-6-hydroxy-2-mercapto-pyrimidine solution for3h,9h,3h or12h, respectively.Cytochrome c can carry out quasi-reversible electrochemical reaction on2-mercapto-pyrimidine,4-amin-2-mercaptopyrimidine,4-hydroxy-2-mercapto-5-methylpyrimidine,4-amino-6-hydroxy-2-mercaptopyrimidine and2-mercaptonicotinic acid modified goldelectrodes, the heterogeneous electron migration rate constant Ks were1.43×10-3cm/s,5.15×10-5cm/s,1.21×10-3cm/s,1.93×10-4cm/s and2.98×10-4cm/s, respectively. The peakpotential difference was61mV,86mV,61mV,67mV and65mV, and the formal potential was19.5mV,17mV,25.5mV,22.5mV and17.5mV, respectively. The peak current ratioipa/ipcwas nearly1. The peak current was positively correlated to the square root of potentialscanning rate (v1/2), indicating that it was a diffusion-controlled quasi-reversible process.3. The effect of molecular structure of modifiers, such as2-mercaptopyrimidine,4-amino-2-mercaptopyrimidine, and4,6-diamino-2-mercaptopyrimidine, on the promotion to the directelectrochemical reaction of cytochrome c were studied by cyclic voltammetry. The promotioneffect to the direct electrochemical reaction of cytochrome c, as well as the stability of modifiedelectrodes, was gradually decreased with the increasing number of amino groups on the ring of2-mercaptopyrimidine, i.e.,2-mercaptopyrimidine>4-amino-2-mercaptopyrimidine, while4,6-diamino-2-mercaptopyrimidine had almost no promotion effect. The positive charges ofmodifiers may be increased with the increasing number of amino groups on the ring of2-mercaptopyrimidine, the electrostatic repulsion between the lysine residues of cytochrome cand the positive charged modifier films was enhanced, the heterogeneous electron migrationrate and the reversibility of the electrochemical reaction of cytochrome c was decreased.4. The promoting effect of4-hydroxy-2-mercapto-5-methylpyrimidine and4-amino-6-hydroxy-2-mercaptopyrimidine modified gold electrodes to the direct electrochemical reactionof cytochrome c were investigated. The promotion effect of modifiers to the directelectrochemical reaction of cytochrome c, as well as the stability of modified electrodes, wasenhanced in the presence of methyl or hydroxyl group on the ring of2-mercaptopyrimidine, andwas decreased in the presence of amino group. It may be caused by the formation of hydrogenbond or electrostatic interaction between hydroxyl of modifies and lysine residues near theelectrochemical active center of cytochrome c.5. The preparation and application of electrochemical biosensor for the determination ofH2O2was also studied, baesd on the catalytic effect of cytochrome c/4-hydroxy-2-mercapto-5-methylpyrimidine/Au (Cyt.c/HMMP/Au), cytochrome c/2-mercaptonicotinic acid/Au (Cyt.c/MNA/Au) electrode to the electrochemical reduction of H2O2.Cytochrome c can be fixed on the surface of4-hydroxy-2-mercapto-5-methylpyrimidineor2-mercaptonicotinic acid modified gold electrodes by hydrogen bond or electrostaticadsorption. A pair of redox peaks of Cyt.c/HMMP/Au or Cyt.c/MNA/Au modified electrodein phosphate buffer solution was observed by cyclic voltammetry, and the peak current waspositively correlated to the potential scanning rate, indicating that it was an adsorption-controlled electrochemical process.The Cyt.c/HMMP/Au or Cyt.c/MNA/Au electrode had good catalytic effect to the electrochemical reduction of H2O2, and can be used as electrochemical biosensor to thedetermination of H2O2with good reproducibility and stability. When determined bychronoamperometry, the peak current was linearly correlated to the concentration of H2O2in therange of2.72×10-4~3.93×10-3mol/L and7.0×10-4~5.47×10-3mol/L, and the detection limit was1.36×10-4mol/L and3.50×10-4mol/L, respectively.