Shenyang Tong

Determination for micro amounts of nucleic acids by a resonance light scattering technique with dequalinium chloride

Based on the strong enhancement effect of nucleic acids on resonance light scattering of dequalinium chloride, the determination method for micro amounts of nucleic acids has been developed. Under the experimental conditions (5.0x10(-5) mol l(-1) dequalinium, pH 7.0, at room temperature) the linear range of this assay is 0.04-10.0 mug ml(-1) for calf thymus DNA and fish sperm DNA, and 0.04-35.0 mug ml(-1) for yeast RNA. The detection limits (3sigma) are 6.2 ng ml(-1) for calf thymus DNA, 7.4 ng ml(-1) for fish sperm DNA, and 7.0 ng ml(-1) for yeast RNA, respectively. Almost no interference can be observed from ionic strength, proteins, nucleoside, and most of the metal ions. Six synthetic samples were determined satisfactorily.

Determination of nucleic acids in acidic medium by enhanced light scattering of large particles

The large particle light scattering technique was first developed as a sensitive and convenient analysis method for microdetermination of nucleic acids by using a common spectrofluorometer. In 0.1 mol l(-1) HCl, H(2)SO(4), or HNO(3) solution, the nucleic acids can aggregate to form large particles whose dimensions are comparable to the wavelength of UV-Vis light. The large particles can result in very strong light scattering which is well proportional to the concentration of nucleic acids in the range of 0.06-100.0 mug ml(-1) for calf thymus DNA, 0.05-60.0 mug ml(-1) for fish sperm DNA, and 0.6-90.0 mug ml(-1) for yeast RNA. The detection limits (3sigma) are 18.0 ng ml(-1) for calf thymus DNA, 16.0 ng ml(-1) for fish sperm DNA, and 57.6 ng ml(-1) for yeast RNA, respectively. Six synthetic samples were determined with satisfactory results.

A linear regression method for the study of the Coomassie brilliant blue protein assay

The interactions of Coomassie brilliant blue G-250 (CBB) with bovine serum albumin (BSA) and gamma-globulin at low pH are investigated by a spectrophotometric method. It is considered that the binding of CBB to protein is because of the weak interactions (ionic, van der Waals, hydrogen bonding, and hydrophobic). The solution equilibria involving the binding of three dye species (blue, green, and red) to protein are treated in the same way as Ringbom model used in the treatment of complexation in analytical chemistry. Based on this treatment, the formation of an isosbestic point in the absorption spectra of CBB-BSA mixtures is discussed, two mathematical models for the description of the CBB protein assay are developed. The first model is a nonlinear equation which is rigorous in theory but unreliable in use because of its optimization procedure. The second model based on an approximation is a linear equation, it allows to estimate apparent binding constant, maximum binding number, and molar absorptivity of bound dye from assay data by a linear regression method. The results of the linear regression operations are reasonable and in agreement with experimental findings. Factors which influence the sensitivity of the CBB protein assay are studied using this method. Ionic strength and acidity are found to have significant effect on the binding of CBB to protein.

The specificity of a chlorphenamine-imprinted polymer and its application.

A chlorphenamine-imprinted polymer was prepared in this study. Chromatographic analysis showed that the retentivity and selectivity of the imprinted polymer were greatly strengthened through molecular imprinting. As a consequence, chlorphenamine could be easily separated from diphenhydramine. Ionic interaction was proved to be the main power for the imprinted polymer to bind chlorphenamine. The strong extraction ability of the imprinted polymer for chlorphenamine in aqueous solution was evaluated further. It was shown that chlorphenamine, as low as 0.02 mumol l(-1), could be concentrated by 50 times with a recovery of more than 90% at pH 5. This study gave the potential of using the imprinted polymer for solid-phase extraction of practical samples.

A novel protein assay method using tetraphenylporphin tetrasulfonate (TPPS4)

Abstract A sensitive protein assay method which involves the reaction of TPPS4 with protein is described. When protein is added to TPPS4 solution, an absorption band with the maximum at 488 nm appears and the absorbance is proportional to the concentration of protein. Just Like the Soret absorption of the porphyrin, the new band is very narrow and there is no overlap at all between them, which means the free dyes would not give any background for the detection of the protein-TPPS4 complexes. A new spectrophotometric method for determination of protein has been constructed and applied to the determination of human plasma protein and urinary protein; The assay using microtiter plates has also been studied.

Nephelometric determination of micro amounts of nucleic acids with protamine sulfate

Nucleic acids can form large particle complexes with protamine sulfate by electrostatic forces, which results in strong light scattering. Based on this, a nephelometric method is described for sensitive and convenient determination of nucleic acids with protamine sulfate by using a common spectrofluorimeter. Maximum light scattering is produced in the range of pH 2.2-4.4 with the same excitation and emission wavelengths at 365 nm. Under optimal conditions, the calibration curves are linear in the range 0.05-60.0 micrograms cm-3 for nucleic acids. The corresponding detection limits are 12.5 ng cm-3 for calf thymus DNA, 9.0 ng cm-3 for fish sperm DNA, and 18.0 ng cm-3 for yeast RNA, respectively. Six synthetic samples are determined with satisfactory results. The relative standard deviation of five replicate measurements is 3.2% for 2.0 micrograms cm-3 calf thymus DNA.

A Study on the Reaction Mechanisms of Protein with BPR-Zn(II) as Spectrum Probe and Its Analytical Application

Abstract The research on the reaction of the metal complex (BPR-Zn2+) as spectrum probe for protein in alkaline medium was conducted in this paper. The experiments indicate that the ternary complex between BPR-Zn2+ complex and protein could be used as a method for protein determination. The reaction conditions, interference materials and sample were studied. A dual-wavelength method is applied to obtain the binding number of spectroscopic probe bound on molecular protein and the reaction mechanism was studied.

Molecular spectroscopic studies on the interaction of glycosaminoglycans with brilliant cresol blue and its analytical application.

The interaction of brilliant cresol blue (BCB) with glycosaminoglycans (GAGs), such as heparin (Hep) and chondroitin 4-sulfate (CS), in aqueous solution has been studied by spectrophotometry and light scattering spectroscopy. Absorbance of BCB at 632 and 594 nm decreased on addition of Hep or CS with the appearance of a new blue-shifted absorption band at 550 nm, which indicated that new metachromatic complex formed. The linear decrease in absorbance of BCB at 632 nm was observed. In addition, Hep was more effective than CS (1.7 times) in decreasing absorbance of BCB. The stoichiometry of Hep or CS with BCB was determined by spectrophotometric titration and the MacIntosh extraction method. The result showed that the stoichiometry of BCB/Hep was 1.8 times that of BCB/CS. These results suggested that the interaction between GAGs and BCB was the result of electrostatic forces, and the differences between Hep and CS were attributed to the different negative charge numbers on repetitive disaccharides unit. Studies on the effects of alcohol and urea indicated that GAGs only interacted with the aggregates of BCB. Moreover, a strong light scattering signal was observed after mixing BCB with GAGs. Furthermore, the light scattering intensity at light scattering bands was proportional to the concentration of Hep or CS added when the concentration of BCB was constant.

Study of the Catalytic Effect of Copper(II)–Protein Complexes on Luminol-H2O2 Chemiluminescence Reaction and its Analytical Application

ABSTRACT Through flow injection analysis experiments, it was discovered that unsaturated complexes of Cu(II) with proteins had a much stronger catalytic effect on the luminol–H2O2 chemiluminescence reaction than did Cu(II). Based on this, a new rapid and sensitive method for determination of some proteins in the range 0.1–20.0 μg ml−1 was established. The detection limits were at the 0.03–0.05μg ml−1 level for bovine serum albumin (BSA), human serum albumin (HSA), human γ–globulin (γ–G), and egg albumin (EA). The method was used for the determination of proteins in human serum with satisfactory results.

Molecular recognition of procainamide-imprinted polymer

A procainamide-specific polymer was prepared by molecular imprinting using methacrylic acid as a functional monomer. It was showed that the imprinted polymer was capable of recognizing the functional difference of amide group and ester group between procainamide and procaine. The procaine-imprinted polymer was also prepared using the same monomer in the same molar ratio of monomer/template, however, it had no such ability. The different molecular imprinting effect of the two molecules showed that changing an amide group for an ester made quite a difference in terms of hydrogen acceptor. The role of the functional groups of the templates in the formation of complementary interacting sites in the polymer and the role of the corresponding interacting sites in the subsequent molecular recognition, was carefully discussed. Accordingly, the mechanism of molecular recognition of this system was proposed. Acrylamide was also used as a functional monomer for the preparation of procainamide-imprinted polymer to be compared with methacrylic acid. It was confirmed that hydrogen bond played an important role in the selectivity of the imprinted polymers.