Posted 13 Dec 2010 22:18
UV - visible absorption spectroscopy for quantitative analysis of a long history, dating back to ancient times, the Year 60 years has been known in ancient Greece Schisandra extract used to estimate the iron content of vinegar, this ancient method is to use the human eye as the initial to testing, so called colorimetry. To the 16,17 century, the relevant theory began to flourish in 1852, Bill (Beer) reference to the cloth to Seoul (Bouguer) 1729 years and Lambert (Lambert) year of articles published in 1760 proposed the basic laws of spectrophotometric , that is equal to the thickness of liquid layer, the color intensity of coloration is proportional to the concentration of the solution, which laid the theoretical basis of spectrophotometry, which is the famous Lambert - Beer law. Photometric instrument also called the formation of spectrophotometry, but somewhat vague concept of spectrophotometry, spectrophotometry is the function of the instrument, that instrument, and by spectrophotometric determination, such instruments include a spectrophotometer and atomic absorption spectrometer (AAS). Spectrophotometry is the nature of light absorption, so called photometric method is more reasonable, of course, said molecular spectrophotometry is most accurate. UV - visible absorption spectra of molecular absorption of the material in the 200-800nm spectral region of light generated. The molecular absorption spectrum resulting from valence electrons and molecular orbitals on the electronic transitions in the electronic energy levels (atoms or molecules in the electronics, is always in a state of a sport. Each state has a certain energy, belongs to a certain level. These e due to various reasons (such as by light, heat, electricity excitation) and from one level to another level, called the transition.) When the electron absorbs the energy from the external radiation a lower energy level transition to a higher energy level. Therefore, each transition corresponds to absorb some energy radiation. With different molecular structure of various substances, have shown selective absorption of electromagnetic radiation characteristics. Spectrophotometry is based on the substance of the selective absorption of electromagnetic radiation characteristics of built up, it belongs to the molecular absorption spectra. The energy absorbed by the transition line with Bohr conditions: Applications UV - visible spectrophotometer can be used for quantitative analysis of materials, structural analysis and quantitative analysis. Determination of some of the compounds but also physical and chemical parameters such as molecular weight, the proportion of complexes with stable Changshu, acid-base ionization constants. 1. Qualitative analysis of compact outside - visible spectrophotometry for the qualitative analysis of inorganic elements is rarely applied, qualitative analysis of inorganic elements can be used atomic emission spectrometry or chemical analysis. Qualitative identification of the organic compounds and structural analysis, due to UV - visible spectroscopy is relatively simple, the characteristic is not strong, so the application of the Act also has certain limitations. But it applies to unsaturated organic compounds. In particular the identification of the conjugated system, to conclude the skeleton structure of unknown compounds. In addition, coupled with infrared spectroscopy, nuclear magnetic resonance spectroscopy and mass spectrometry for qualitative identification and structural analysis, it is nonetheless a useful adjunct. There are two general methods of qualitative analysis, comparison with the absorption curve and the rule of thumb for calculating the maximum absorption wavelength of the max, and then compared with the measured value. 2. Structural analysis structural analysis of compounds can be used to determine the configuration and conformation. Isomers, such as identification and tautomers. 3. Quantitative analysis of UV - VIS spectrophotometer is based on quantitative analysis of Lambert-Beer law, which was measured in a certain wavelength and its physical solubility of absorbance was linear. In response to this, the solution by measuring the absorbance of certain wavelengths of incident light can be obtained in the solution of the substance concentration and content. The determination of commonly used methods are: single-component quantitative method, multi-component quantitative method, dual-wavelength method, and the differential spectrophotometric derivative spectrometry. 4. Complex composition and stability constants of complexes composed of commonly used measure in two ways: the molar ratio method (also known as saturation method) and the continuous changes in equimolar method (also known as the Job method). 5. Acid-base spectrophotometric determination of dissociation constants is to measure the application of analytical chemistry reagent indicator or the dissociation constant of the common methods, the method is particularly suitable for low solubility of the weak acid or weak base. Category UV - visible spectrophotometer type a lot, but can be summarized into three types: single-beam spectrophotometer, double beam spectrophotometer and dual wavelength spectrophotometer. Single-beam spectrophotometer the optical path diagram as the previous figure (UV - visible spectrophotometer basic structure of the graph) shows, the monochromatic parallel light beam after splitting, in turn through the reference solution and sample solution tuck, for Determination of absorbance. This type of spectrophotometer simple structure, convenient operation, easy maintenance, suitable for routine analysis. Domestic 722, 751 type, 724 type, the British-based SP500 and the BackmanDU-8 type Dengjun belong to this category photometer. Double-beam spectrophotometer the optical signal as shown below, after splitting by the monochromator by mirror (M1) is divided into two beams of equal curvature, a bunch of pool by reference, and the other a bunch through the sample cell. Photometer can automatically compare two beams of light intensity, this ratio is the transmittance of the sample, after logarithmic transformation to convert it as a function of wavelength absorbance recorded. Double beam spectrophotometer in general can automatically record the absorption spectrum curve. Since the two beams, respectively, while the reference cell and sample through him, the light intensity changes can automatically eliminate the errors caused. Such instruments have made 710, 730 type and 740 type, Hitachi 220 series, Shimadzu, -210, British UNICAMSP-700 and so on. The basic dual-wavelength spectrophotometer light path as shown below. The light emitted by the same light source is split into two beams, respectively, after the two monochromator to obtain two beams of different wavelengths of monochromatic light; re-use allows the two beams of light cut with some frequency alternating light the same absorption cell, and then through the optical photomultiplier tubes and electronic control systems, and finally by the display shows the absorbance difference between the two wavelengths A (A = A1-A2). The advantages of dual-wavelength spectrophotometer: For multi-component mixture, turbidity samples (such as biological tissue fluid) analysis, and the presence of background interference or coexistence of components in the absorption of interference, the use of dual wavelength spectrophotometry, often improving The sensitivity and selectivity. Using dual-wavelength spectrophotometer, can be derivative spectra. By converting optical system, so dual-wavelength spectrophotometer can be easily converted into single-wavelength work. If the two wavelengths were recorded in absorbance versus time curve, but also for chemical reaction dynamics.