| This thesis is maily focused on intensive research on the recognition and detection of certain biological molecules and drug molecules based on the modified electrode and molecular imprinting technique using new nano-composite materials. The thesis is divided into two parts:one is the determination of small biological molecules and drug molecule by modified electrode; another is the identification of protein macromolecules by molecularly imprinted materials.Ascorbic acid (AA), dopamine (DA), uric acid (UA) and tryptophan (TRP) are considered as vital small biomolecules for physiological processes in human metabolism. Usually they coexist in biological matrixes. Abnormal levels of these species will lead to several diseases and disorders. Propranolol is one of a number of β-adrenergic antagonists (β-blockers) used extensively in the treatment of cardiovascular disorders such as hypertension and angina pectoris. So it is very important to develop a simple and effective method for the determination of these small molecules in the field of biomedical chemistry and diagnostic research. Electrochemical technique arouses grate interest for its rapid response, high sensitivity, simple operation and low cost. However, at conventional electrodes, the oxidation potentials of these analysts are close enough with a pronounced fouling effect, resulting in poor selectivity and reproducibility. To overcome these problems, developing new nano-composite materials to modify electrodes has a very important theoretical and practical significance.The left part of the thesis enclosed identification of biological macromolecules on the basis of the molecular imprinting technique (MIT). MIT is an attractive method for the generation of polymer-based molecular recognition elements tailor-made for a given target or group of target molecules. We presented a novel and simple method to create magnetic inprinting microspheres (MIMs) for the protein. MIMs combined magnetic microspheres as separation tool and silica shells as biocompatible recognition interface. This dissertation includes six chapters as follows:In chapter one, we pointed out the theoretical and practical significance of this thesis, introduced the basic principle, applications and development prospects of new nano-materials, chemically modified electrodes and molecular imprinting technique.Then we proposed the research scheme of the recognition and determination of biological molecules and drug molecules. Finally we outlined the experimental ideas and the research purpose.In chapter two, a sensitive molecularly imprinted electrochemical sensor was created for selective detection of propranolol hydrochloride using dopamine as monomer by the method of electropolymerization on glassy Carbon Electrode (GCE) modified with multi-walled carbon nanotubes (MWNTs). The electrochemical performance and morphology of the imprinted sensor were characterized by scanning electron microscope (SEM), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Test conditions were optimized and the selective response to the structurally similar compounds of propranolol hydrochloride was studied. The results showed that the sensor had good selectivity and the sensitivity increased greatly due to the introduction of MWNTs. A linear relationship was obtained between the current and the concentration of propranolol hydrochloride over a range from 0.20 to 100 umol/L, with a detection limit of 2.53×10-8 mol/L (S/N=3). Moreover, the sensor exhibited excellent stability and reproducibility. The method was applied to detect propranolol hydrochloride in drug tablets and the recovery ratio was 97.3-104.0%. In addition, the accuracy of this method was examined by comparison with the result of the Chinese Pharmacopoeia method. It means that the method proposed could be reliably used for routine analysis.In chapter three, Preparation and application of magnetic molecularly imprinted polymers. The amino magnetic particles with a diameter of about 150 nm were prepared by one-step method first, and reacted with glutaraldehyde to obtain the matrix material for fixing protein molecules. UV, IR and XPS were employed to characterize the materials. The aluminosilicate gel was used to polymerize on the surface of magnetic microspheres, and followed with elution carried out in the solution of oxalic acid. But the result showed that there was no polymer wrapping the magnetic particles. In view of this result, in the first step, magnetic particles were coated by silane, and silane was used as monomer and crosslinker to prepare the molecularly imprinted polymers for protein. The amount of silane and the pH in the reaction with glutaraldehyde were optimized. The amount of protein fixed on the surface increased. The re-adsorption experiments and preliminary competition experiments were carried out.In chapter four, Preliminary identification and detection of the protein molecules were explored by combining microfluidic chip with molecular imprinting technique. The detailed routine as follows:Firstly, molecular imprinted membrane was prepared in the chip channel. Then isolation was introduced to occupy vacant binding cavities. The channel was then incubated in a horseradish peroxidase-labeled protein solution. At last, the target molecules in sample replaced the enzyme-labeled molecules, and the detection realized through the current changes.In chapter five, Multi-walled carbon nanotubes decorated flower-liked graphene nanocomposite for the simultaneous determination of AA, DA, UA and TRP was obtained facilely. First, by using zeolite Ni-MCM-22 as a catalyst and template, single graphene sheets (TGS) can be easily synthesized which has a large specific surface area and high conductivity than graphene sheets (RGS) synthesized by chemically reduction. When TGS were mixed with MWNTs under ultrasonication, MWNTs/TGS nanocomposites were formed via π-π stacking interaction. Simultaneous determination of AA, DA, UA and TRP was achieved on MWNTs/TGS modified GCE. Compared with MWNTs/GCE, TGS/GCE and MWNTs/RGS/GCE, MWNTs/TGS/GCE shows a higher catalytic activity and better selectivity. The electrochemical performance and morphology of the modified electrode were characterized by SEM, CV and DPV. Under the selected conditions, the calibration curves for AA and DA were obtained in the range of 0.1 mM-6mM,0.3μmol/L～10 μmol/L with detection limits (S/N=3) of 18.28 μmol/L and 0.06 μmol/L. For UA, two linear calibrations were obtained in the range of 5 μmo1/L-100 μmol/L and 0.3 mmol/L～1 mmol/L with detection limits (S/N=3) of 0.93 μmol/L. For TRP, there were also two linear calibrations in the range of 5 μmol/L～30 μmol/L and 60μmo1/L-500 μmol/L with detection limits (S/N=3) of 0.87 μmol/L (S/N= 3). Moreover, the modified electrode exhibited excellent stability and reproducibility and could be used in simultaneous determination of AA, DA, U A and TRP in real samples.In chapter six, we summarized and gave objective evaluations of the research results achieved. Meanwhile, the problems and inadequacies exist in the study were pointed out and we proposed the subsequent objectives and research ideas. |
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