| Postgraduate: Liu Dianmei Tutor: Pro. Zhao ShulinProtein is one of the most important materials in the life body, and is the base of life. Quantitative analysis of protein is often concerned with biochemistry, pharmacy, other branches of biochemistry and life sciences, is the routine analytical item in clinical assay and food survey. Serum albumin is the protein which possesses the wide-link ability, could be linked to large, medium, and small-sized molecules. Meantime Serum albumin is also the main ingredient which can form blood glue osmotic pressure. It plays an important role in maintaining the normal distribution of body fluid and keeping blood content and blood pressure, etc. So it is of great significance to study the interaction of protein with mental complex and establish new protein assay methods, especially for disease diagnosis and life sciences research.In this paper, based on the special mental complex' development, we probed into the conditions they form supermolecula complex with proteins, studied the supermolecula complex's absorption spectral behaviors and the mechanism protein react on mental complex, developed a series of new type spectroscopic probe for proteins, thus four novel methods to assay proteins were established.The dissertation here is divided into four parts.In part one , a fading reaction of barium(Ⅱ)-chlorophosphonazo Ⅲ (Ba(Ⅱ)-CPAⅢ)with protein was studied in detail. And the effects of some materials such as amino acid, urea, mental ions etc. were also checked. The optimal conditions and the absorption spectra character were studied. The absorbance strength descend linearly after react with the protein. Based on it anovel spectrophotometric method for the determination of protein was established. The apparent molar absorptivity of the binding reaction is 6.86×105 L·mol-1·cm-1 .The method can be used to determine 0~40 mg/L of protein. The method is sensitive and selective and it has been applied to the detection of total amounts of proteins in human serum samples successfully. It was considered that the reaction mechanism of protein with Ba(Ⅱ)-CPAⅢ complex was similar to that of cationic surfactant, and the electrostatic force is the main binding force In part two, the interaction between protein(bovine serum albumin) and barium(Ⅱ)-p-acetyl-chlorophosphonazo(Ba(Ⅱ)-CPA-pA) was investigated and a simple sensitive and rapid protein assay method based on it was established. At pH 2.0~2.8 and in the presence of ethanol and triton X-100, the binding reaction was complete rapidly, and caused the absorbance decrease at 649 nm of Ba(Ⅱ)-CPA-pA complex. The molar absorptivity of the binding reaction is 1.56×106 L·mol-1·cm-1. The linear range of standard curve is 0~40 mg/L for BSA. Results for human serum album agreed well with those obtained by Pyrocatechol Violet(PV)-Molybdenum(Ⅵ) method.In part three, the absorption spectral behavior of binding reaction of cuprum(Ⅱ)- p-nitro-chlorophosphonazo (Cu(Ⅱ)-CPA-pN) complex on bovine serum albumin(BSA) was investigated. In acidic solution, the color of Cu(Ⅱ)-CPA-pN is blue while Cu(Ⅱ)-CPA-pN-BSA complex is purple. A new method for the determination of protein by using Cu(Ⅱ)-CPA-pN complex as a spectroprobe was developed. The method is simple, rapid, stable and can be used to assay 0-40 mg/L of protein. The apparent molar absorptivity of the binding reaction is 5.43×105 L·mol-1·cm-1. The binding number of BSA with the complex was estimated by molar ratio method and Chial method with comparison. It was proved that there were two sorts of binding sites in BSA reacting with the Cu(Ⅱ)-CPA-pN complex.In part four, for the first time, a novel sensitive method is described for the Spectrophotometric determination of protein. Based on the inhibitinginteraction of protein to barium(Ⅱ)- p-nitro-chlorophosphonazo (Ba(Ⅱ)-CPA- pN) β-complex. In definite conditions, the reaction between Ba(Ⅱ) and CPA-pN is β type, there is a strong absorbance at 736 nm ,and small amounts of protein...
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