Study On Some New Methods For Determination Of The Nucleic Acid And Medicines By Molecular Spectrometry

Molecular spectroscopy is established by determining the spectral characteristics(intensity and wavelength in the reflection, absorption or emission, scattered radiation), which is applied for the qualitative analysis, quantitative analysis and structure analysis. Molecular spectroscopy is the basis of development and experiment of molecular orbital theory. It is the important experimental method of determining and indentifing molecular structure. According to difference ranges of absorbing the electromagnetic wave, it can be divided into the near-infrared spectrum, infrared spectrum and ultraviolet-visible spectrum etc and so on; according to the difference electromagnetic wave emission method, it can be divided into the molecular fluorescence, the phosphorescence and the scattering spectrum etc. In this paper, we have studied and developed the new stystem and method that Vindensine-erythrosine, Mercaptopurine and DNA were determining by the molecular spectroscopy. we mainly investigated the spectrophotometric, fluorescence and resonance Rayleigh scattering spectral characteristics that the interaction between drugs/DNA and metal ions or halofluorescein. Besides, the optimum conditions, the influencing factors and their analytical applications were investigated. In addition, we discussed the reaction mechanism for change of RRS spectrum.Supported by the Project Foundation of Chongqing Municipal Education Committee(KJ081306), this article studies on some new applications of Molecular spectroscopy in analytical chemistry:1. Study on the determination of VDS by UV, FL, RRS, and the interaction between VDS and PW, in order to improve sensitivity of RRS spectral analysis and expand its analytical application; 2. Study on the new application of RRS spectrum method in the medicine analysis, especially in the investigation of the interaction between metallic ion with Mercaptopurine and Mercaptopurine-metal chelate with nucleic acids, and developed new methods for the determination of Mercaptopurine, nucleic acids by RRS method. 1. Study on the Interaction of Vindensine with Halo-fluorochrome by Molecular Spectrometry and their Analytical Applications1.1 The absorption, fluorescence and resonance Rayleigh scattering spectra of vindensine-erythrosine ion-association complex system and their analytical applicationsIn pH 2.7 Britton-Robinson (BR) buffer solution, erythrosine (Ery) reacts with Vindensine sulfate (VDS) could form ion-association it results in the change of absorption spectra, the quenching of fluorescence and the great enhancement of resonance Rayleigh scattering (RRS). The absorption of wavelength is located at 520 nm. The maximum excitation (λex) and emission (λem) wavelengths are at 523 nm and 551 nm, respectively. The maximum RRS wavelength is located at 325 nm. The increment of absorption, the quenching of fluorescence and the enhancement of RRS are all proportional to the VDS concentration in a certain range. The quantitative determination ranges and the detection limits of the three methods for VDS are 0.8～4.0μg·mL-1 and 0.120μg·mL-1 (spectrophotometry),0.2～3.0μg·mL-1 and 0.072μg·mL-1 (fluorimetry),0.08-2.0μg·mL-1 and 0.019μg·mL-1 (RRS), respectively. Based on Ery, the sensitive method for the determination of VDS have been developed. The RRS method could be applied to the determination of VDS in urine samples with satisfactory results. The mechanisms of the RRS enhancement were discussed.1.2 Fading Spectrophotometric Determination of Vindensine-erythrosine with RBIn pH 2.7 Britton-Robinson (BR) buffer solution, the interaction of Vindensine sulfate (VDS) with RB resulted in the change of absorption spectra. The maximum fading wavelength was located at 526 nm. The fading degree and the concentration of VDS were followed by the law of Lambert-Beer's in the quantitative determination ranges of 0.5～6.0μg ml-1, and the detection limit was 0.29 ug ml-1. In this work, the characteristics of the absorption spectra of fading reaction, the optimum reaction conditions and the effects of foreign substances were investigated, and the results indicated that the proposed method had a good selectivity. The spectrophotometric method was applied for the determination of VDS in urine samples. 2. Resonance Rayleigh scattering spectra and resonance non-linear scattering of 12-tungstophosphoric acid-Vindensine sulfate system and its analytical applicationIn 0.2 mol/L的HCl medium,12-tungstophosphoric acid (PW) reacts with Vindensine sulfate (VDS) could form ion-association, which lead to the great enhancement of resonance Rayleigh scattering (RRS) and second-order scattering (SOS) and frequency doubling scattering (FDS). The maximum scattering wavelengths were 308 nm for RRS,564 nm for SOS and 390 nm for FDS, respectively. The increments of scattered intensity (△I) of products were directly proportional to the concentrations of VDS in certain ranges. The linear ranges and detection limits of VDS were 0.08-2.0μg/mL and 7.0 ng/mL(RRS method),0.08-2.5μg/mL and 14 ng/mL (SOS method), and 0.08-2.5μg/mL and 11 ng/mL (FDS method). Taking the RRS method of highest sensitivity as an example, the optimum reaction conditions of RRS method and the impact factor were investigated. The effects of foreign substances were examined and it showed that the method had a good selectivity. Based on the above phenomena,a highly sensitive, simple and fast method for the determination of VDS has been developed. RRS method was applied for the determination of VDS in serum and urine samples.3. Resonance Rayleigh Scattering Spectra of Pd(Ⅱ) and Mercaptopurine Intractive and their Analytical ApplicationsIn pH 4.8 B-R buffer solution, The metal ion Pd(Ⅱ) could react with Mercaptopurine(MP) to form stable coordination compound, theirs moles ratio was 1:1. At the same time the resonance Rayleigh scattering (RRS) spectra were been enhancemented and appeared new and wide RRS peak, we selected the highest peak located at 400 nm as detection wavelength. Because there was a linear relationship between the RRS enhanced intensity (△I) and the MP concentration, and the linear range were 0.05～3.6μg·mL-1 and detection limit (3σ) was 17.0 ng·mL-1, So a new method for determination of the MP can be developed. The method had fine selectivity and had been applied to determination of the MP in serum and urine sample with satisfaction result.4. Study on Resonance Rayleigh Scattering Spectra of Cu(Ⅱ)-Mercaptopurine and Nucleic Acid SystemIn pH 4.6-4.9 B-R buffer solution, Mercaptopurine(MP) could react with Cu(Ⅱ) to form stable chelate compound, theirs resonance Rayleigh scattering (RRS) spectra were been enhancemented and appeared new RRS spectra and the highest peak located at 453 nm as detection wavelength. Thus added herring milt DNA(hsDNA), Salmon milt DNA(sDNA) and calf thymus DNA(ctDNA) into the binary system could be lead to its RRS spectra quenching, and the weaken degree of spectrum was ratio to the concentration of DNA. The paper investigated the absorption and RRS spectra of reaction products and eligible reaction conditions and influencing factor. So a new method for determination of DNA can be developed and succeed to applied to determination of the DNA artificial synthetize samples.