Chapter 26 Molecular Absorption Spectrometry
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1 Chapter 26 Molecular Absorption Spectrometry Molecular spectroscopic methods are among the most widely used of all instrumental analytical methods. Molecular spectroscopy is used for the identification and determination of a huge number of inorganic, organic and biochemical species. Molecular ultraviolet/visible absorption spectroscopy is employed primarily for quantitative analysis. Infrared absorption spectroscopy is one of the most powerful tools for determining the structure of both inorganic and organic compounds. 1 ULTRAVIOLET/VISIBLE MOLECULAR ABSORTION SPECTROSCOPY In the UV/visible region, many types of inorganic compounds absorb radiation directly. Others can be converted to absorbing species by means of a chemical reaction. Absorption measurements in the UV/visible region of the spectrum provide qualitative and quantitative information about organic, inorganic, and biochemical molecules. 2 Absorption by Organic Compounds: Two types of electrons are responsible for the absorption of ultraviolet and visible radiation by organic molecules: (1) shared electrons that participate directly in bond formation and (2) unshared outer electrons that are largely localized on atoms such as oxygen, the halogens, sulfur, and nitrogen. The shared electrons in single bonds are so firmly held that absorption occurs only with photons more energetic than normal UV photons. Electrons involved in double and triple bonds of organic molecules are more loosely held and are therefore more easily excited than electrons in single bonds. Thus, species with unsaturated bonds generally absorb in the UV. Unsaturated organic functional groups that absorb in the UV/visible region are 3 4 known as chromophores.
2 5 6 Absorption by Inorganic species: In general, the ions and complexes of elements in the first two transition series absorb broad bands of visible radiation in at least one of their oxidation states and are, as a consequence, colored. Absorption involves transitions between filled and unfilled d- orbitals of the metal ion with energies that depend on the bonded ligands. The energy differences between these d-orbitals and thus the position of the corresponding absorption maximum depend on the position of the element in the periodic table, its oxidation state, and the nature of the ligand bonded to it. 7 8
3 Charge-Transfer Absorption: For quantitative purposes, charge-transfer absorption is particularly important because molar absorptivities are unusually large, a circumstance that leads to high sensitivity. Many inorganic and organic complexes exhibit this type of absorption and are therefore called charge-transfer complexes. A charge-transfer complex consists of an electron-donor group bonded to an electron acceptor. When this product absorbs radiation, and electron from the donor is transferred to an orbital that is largely associated with the acceptor. The excited state is thus the product of a kind of internal oxidation/reduction process Qualitative Analysis Qualitative applications of UV/visible spectroscopy are limited because the spectra of most compounds in solution consist of one or, at most, a few broad bands with no fine structure that would be desirable for unambiguous identification. The spectral position of an absorption band is, however, an indication of the presence or absence of certain structural features or functional groups in a molecule. Usually, UV/visible absorption spectroscopy is only used for confirmation in conjunction with a more useful qualitative technique, such as NMR, IR, and mass spectrometry
4 Fundamental Studies Spectrophotometry in the UV/visible region is one of the major tools for studying chemical equilibria and kinetics. Wavelengths are chosen to allow monitoring of one or more reactants, products, or intermediate species. The concentrations are then obtained by using Beer s law with known or previously determined molar absorptivities. A wide variety of reaction types have been studied in this way. From Beer s law, the final concentrations of reactants and products are obtained and equilibrium constants determined from known stoichiometric relationships. In kinetic studies, spectrophotometry is used to monitor the appearance of a product or intermediate, or the disappearance of a reactant. 13 Quantitative Analysis Ultravilet/visible spectrophotometry is one of the most powerful and widely used tools for quantitative analysis. Important characteristics of UV/visible spectrophotometry include wide applicability to organic, inorganic, and biochemical systems; good sensitivity; detection limits of 10-4 to 10-7 M; moderate to high selectivity; reasonable accuracy and precision (relative errors in the 1 to 3% range and with special techniques, as low as a few tenths of a percent); and speed and convenience. In addition, spectrophotometric methods are readily automated. 14 Standards and the Calibration Curve: In most spectrophotometric methods, calibration is achieved by the method of external standards. Here, a series of standard solutions of the analyte is used to construct a calibration curve of absorbance versus concentration or to produce a linear regression equation. The slope of the calibration curve or regression equation is the product of absorptivity and pathlength. Thus, using external standards is way of determining the proportionality factor between absorbance and concentration under the same conditions and with the same instrument as is used for the samples. 15 The Standard Addition Method: The difficulties associated with production of standards with an overall composition closely resembling that of the sample can be formidable. Under such circumstances, the method of standard additions may prove useful. In the single-point standard addition method, a known amount of analyte is introduced into a second aliquot of the sample and the difference in absorbance is used to calculate the analyte concentration of the sample. Alternatively, multiple additions can be made to several aliquots of the sample and multiple standard addition calibration curve obtained. 16
5 17 Analysis of Mixtures: The total absorbance of a solution at any given wavelength is equal to the sum of the absorbances of the individual components in the solution. This relationship makes it possible in principle to determine the concentration of the individual components in a mixture even if there is strong overlap in their spectra. There is no wavelength at which the absorbance is due to just one of these components. To analyze the mixture, molar absorptivities are first determined at wavelengths λ 1 and λ continued The wavelengths selected are ones at which the two spectra differ significantly. Thus, at λ 1, the molar absorptivity of component M is much larger than that for component N. The reverse is true for λ 2. To complete the analysis, the absorbance of the mixture is determined at the same two wavelengths. From the known molar absorptivities and pathlength, the following equations hold: A 1 = ε M1 bc M + ε N1 bc N A 2 = ε M2 bc M + ε N2 bc N 19 20
6 Spectrophotometric Titrations Ultraviolet/visible spectrophotometric and photometric measurements are useful for locating the end points of titrations. The method requires that one or more of the reactants or products absorb radiation or that an absorbing indicator be present. In spectrophotometric titrations, the spectrophotometer serves as the detector that monitors the transmittance or absorbance or the solution at a suitable wavelength during the addition of increments or the titrant. Acid/base titration can be monitored spectrophoto-metrically by adding a small amount of an indicator that is colored in either the acidic or basic form. 21 Titration Curves: The plot of absorbance versus titrant volume is called a spectrophotometric titration curve; the shapes depend on the species that absorbs radiation. Normally, the absorbances are corrected for dilution by the titrant by multiplying the measured values by (V T + V A )/V A, where V A and V T are the volumes of the analyte solution and titrant, respectively. Ideally, the end point is located by a sharp change in absorbance; often, conditions are arranged so that two straight-line regions of differing slopes intersect at the end point. If the reaction is not quantitative near the equivalence point, the linear segments before and after the end point can be extrapolated to locate the end pint; adherence to Beer s law is a necessity Applications of Spectrophotometric Titrations: Spectrophotometric or photometric titrations have been applied to many types of reactions. Most standard oxidizing agents have characteristic absorption spectra and thus produce photometrically detectable end points. Although standard acids or bases do not absorb, the introduction of acid/base indicators permits the use of spectophotometric end points in neutralization titration. The photometric end point has also been used to great advantage in titration with EDTA and other complexing agents. 24
7 25 continued At 745 nm, the cations, the reagent, and the bismuth complex formed in the first part of the titration do not absorb, but the copper complex does. Thus, the solution exhibits no absorbance until essentially all the Bi(III) has been titrated. With the first formation of the Cu(II) complex, an increase in absorbance occurs. The increase continues until the copper end point is reached. Additional reagent causes no further absorbance change. Clearly, two well-defined end points result. 26 INFRARED ABSORPTION SPECTROSCOPY Infrared absorption spectroscopy is also widely employed in analytical chemistry for identification. Its scope is nearly as broad as that of UV/visible methods. In the IR region, absorption of radiation can give information about the identity of compounds, the presence or absence of functional groups, and the structure of molecules. IR absorption is one of the premier techniques for qualitative analysis and functional group identification. 27 Molecules That Absorb Infrared Radiation With the exception of homonuclear diatomic molecules, such as O 2, Cl 2, and N 2, all molecules, organic and inorganic, absorb infrared radiation. Absorption of IR radiation involves transitions among the vibrational energy levels of the lowest excited electronic energy levels of molecules. The number of ways a molecule can vibrate is related to the number of bonds it contains and thus the number of atoms making up the molecule. The number of vibrations is large even for a simple molecule. 28
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