| The quantitative analysis of nucleic acid and protein is of great importance in biochemistry and clinic medicine. The common analytical methods include spectrophotometry and fluorimetry, but they have some serious disadvantages. Spectrophotometry suffers from low sensitivity, multiple steps, and long time; fluorimetry is restricted by fluorimetric systems and fluorimetric reagents are toxic and expensive. Resonance light scattering (RLS) technique is a new analytical method with high sensitivity, simplicity, rapidity, and good selectivity.Small molecule combines with nucleic acid by three modes: intercalative binding, groove binding and electrostatic force. Such research helps to discover the mechanism of pathological change and develop new anti-cancer medicine, so it has great significance.In this dissertation six new RLS methods were established and the binding mechanism of small molecule and nucleic acid was discussed.Firstly, tetraphenyl porphyrin cobalt chlorine (CoTPPCl) was utilized to determinate nucleic acid. Nucleic acid enhances the weak RLS signal of CoTPPCl under the optimum conditions, and the enhanced light scattering intensity is proportional to the concentration of nucleic acid in the range of 0.05—3.5 μg ml-1 for calf thymus DNA, 0.03—4.2 μg ml-1 for fish sperm DNA and 0.02—4.9 μg ml-1 for yeast RNA. The detection limits are 3.5 ng ml-1 for calf thymus DNA, 4.5 ng ml-1 for fish sperm DNA and 6.1 ng ml-1 for yeast RNA, respectively. Synthetic samples were determined with satisfactory results.Secondly, a new RLS method for the determination of nucleic acid was developed by using tetraphenyl porphyrinatoiron chlorine (FeTPPCl). FeTPPCl binds with nucleic acid by intercalation and aggregates on the surface of nucleic acid to enhance RLS signals. The proposed method is more sensitive than that of CoTPPCl.Thirdly, μ-oxotetraphenyl porphyrinatoiron [(FeTPP)2O] was employed to determinate nucleic acid. Besides coordinating with nitrogen atoms on the porphyrin cycle, the iron ion combines with the oxygen atom, thus the radii of the iron ion reduces so it can coordinate with the porphyrin cycle in the same plane. The rigidity and coplanarity of the molecule are improvedgreatly, which is profitable to the RLS enhancement. The linear ranges are 0.02—2.5 ng ml"* f°r calf thymus DNA, 0.02—2.7 ug ml"1 for fish sperm DNA, and 0.03—3.1 ug ml"1 for yeast RNA. The detection limits (3a) are 1.0 ng ml"' for calf thymus DNA, 1.1 ng ml"1 for fish sperm DNA and 1.2 ng ml"1 for yeast RNA, respectively. In comparison with mononuclear metalloporphyrin, binuclear metalloporphyrin is more sensitive.Fourthly, zwitterionics cocamidopropyl hydroxysultaine (HSB) was firstly used to determinate DNA. HSB has one cationic group and several anionic groups. After nucleic acid is added, the nitrogen atom of HSB combines with the negative phosphate groups of nucleic acid. Apart from this, the anionic group of zwitterionics binds with the positive bases of nucleic acid. The linear range is 0.02—4.25 ug ml"1 and the detection limit (3a) is 1.5 ng ml"1 for calf thymus DNA. Compared with those of cationic surfactants, the proposed method has a wider linear range and lower detection limit.Fifthly, a novel assay of proteins was established based on enhanced RLS signal of ammonium molybdate. Under optimum conditions ammonium molybdate reacts with proteins to form a new supermolecule complex. The molecule weight enhancement of scattering particles and the hydrophobicity of supermolecule complex enhance RLS signal. The enhanced RLS intensity is proportional to the concentration of proteins in the range of 0.25—8.5 p.g ml"1 for bovine serum albumin and 0.40—10.2 jig ml"1 for human serum albumin. The detection limits (3fr) are 12.5 ng ml"1 for bovine serum albumin and 14.4 ng ml"1 for human serum albumin. The results of the determination for human urine and serum samples are close to those obtained from the Coomassie Brilliant Blue method.Sixthly, phosphotungstic acid was employed to determinate proteins in the presence ofdodecylbenzenesulfonate sodium by the RLS technique. Phosphotungstic acid combines with protein, and then dodecylbenzenesulfonate sodium forms a micelle on the surface of phosphotungstic acid-protein. It reduces the distance of phosphotungstic acid and protein, and enlarges the volume of scattering particles. The formation of micelle is helpful for the RLS enhancement. The linear ranges of bovine serum albumin and human serum albumin are 0.07— 4.5 p.g ml"1 and 0.08—4.9 ^g ml"1. The detection limits (3&) are 6.8 ng ml"1 for bovine serum albumin and 7.9 ng ml"1 for human serum albumin. The proposed method is more sensitive than that of ammonium molybdate.The chief characteristics of this thesis are as follows:1. We studied three metalloporphyrin-nucleic acid systems, analyzed their difference and established three new assays of nucleic acid. We firstly used metalloporphyrins to determinate nucleic acid and found that there exists a rule for their sensitivities: binuclear metalloporphyrin >mononuclear metalloporphyrin >nonmetal porphyrin. It can be concluded that coplanarity of RLS probes is a key factor of sensitivities. Three metalloporphyrins bind with nucleic acid in the modes of intercalation, which offers a convictive proof of filtrating anti-cancer medicine.2. Zwitterionics was firstly performed to determinate nucleic acid. In our opinions, anionic and cationic groups of zwitterionics bind with phosphate groups and bases of nucleic acid to produce a large-sized complex by electrostatic force, which enhances RLS signals. The proposed method is more sensitive than cationic surfactants. The work will be published in Analytica Chimica Acta.3. We investigated two poly acid-protein systems, discussed mechanism of RLS enhancement, and proposed two new methods for the determination of protein. The common protein probes are primarily anionic dyes, but ammonium molybdate and phosphotungstic acid enlarged the range of RLS probes. For the phosphotungstic acid method, the utilization of dodecylbenzenesulfonate sodium forms a micelle and improves the sensitivity of the method.
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