Studies On Two Kinds Of Novel Electrochemical Biosensors Based On Hemoglobin

As the main protein in red blood cells of human, hemoglobin is the basic substance that transports oxygen in blood. It is widely used as a model to study the direct electrochemistry and biosensing of redox proteins because of its similar properties as enzymes, known and documented structure, and commercial availability. Much attention has been devoted to the development of the third-generation electrochemical biosensor based on the direct electrochemistry of redox proteins. Therefor, in this thesis, we fabricated and developed the novel third-generation electrochemical biosensors based on the direct electron transfer between hemoglobin and the underlying electrode, and also explored the direct electrochemistry of hemoglobin and its resulting biosensors. The main points are summarized as follows:1. Hemoglobin (Hb) modified with mercaptopropionic acid-stabilized CdTe quantum dots (QDs) was immobilized on a glassy carbon electrode by chitosan. QDs could accelerate the electron transfer between Hb and the electrode, as well as its catalytic ability toward nitrite (NO2-). In aqueous-ethanol (V/V=1) mixtures, the modified electrode displayed a couple of stable and well-defined redox peaks, and the redox peak currents increased linearly with an increasing scan rate ranging from50to500mV/s, indicating a surface-controlled electrochemical behavior, with an electron transfer rate constant (ks)1.03s-1, at the scan rate of100mV/s. UV-Vis spectroscopy analysis showed that the modified Hb could keep its natural structure. The modified electrode showed an electrocatalytic activity to the reduction of NO2-without the aid of an electron mediator, with a smaller apparent Michaelis-Menten constant (Kmapp)0.202mmol/L. The electrocatalytic response showed a linear dependence on the NO2-concentration ranging from7.5to120.0μmol/L with a fairly low detection limit of2.5μmol/L (S/N=3).2. Hb-PDDA films were constructed by encapsulating hemoglobin in PDDA matrix and modified on the glassy carbon electrode (GCE), where PDDA is a polycation poly(diallyldimethylammonium chloride), realizing the direct electrochemistry between Hb and the electrode surface without any nanomaterial. UV-Vis absorption testified Hb could still keep its native structure in Hb-PDDA films. The influence of the ethanol proportion to the electrochemistry of Hb-PDDA was investigated in PBS-ethanol mixtures by cyclic voltammetry (CV). CV of Hb-PDDA/GCE showed a pair of reversible and stable peaks in PBS-ethanol solution (V/V=1, pH7.0) at the scan rate of100mV/s, with an electron transfer rate constant (ks)1.03s-1. Hb-PDDA/GCE also achieved the electrocatalytic reduction to H2O2in microenvironment of the PDDA films at-0.420V (vs. SCE), with a smaller apparent Michaelis-Menten constant (Kmapp)0.496mmol/L and a lower detection limit0.9μmol/L.