1. Chapter 26 Molecular Absorption Spectrometry

2. 26 A Ultraviolet and visible molecular absorption spectroscopy Ultraviolet and visible radiation absorption by molecular species can be used for qualitative and quantitative analyses. UV-visible absorption is used to monitor titrations and to study the composition of complex ions. Absorption generally occurs in one or more electronic absorption bands, each of which is made up of many closely packed but discrete lines. Each line arises from the transition of an electron from the ground state to one of the many vibrational and rotational energy states associated with each excited electronic energy state.

3. Absorption of radiation by organic molecules in the wavelength region between 180 and 780 nm results from interactions between photons and electrons that either participate directly in bond formation or that are localized about atoms. The wavelength of absorption of an organic molecule depends on how tightly its electrons are bound. The shared electrons in carbon-carbon or carbon-hydrogen single bonds are so firmly held that their excitation requires energies corresponding to wavelengths in the vacuum ultraviolet region below 180 nm. Electrons in double and triple bonds of organic molecules are not as strongly held and are therefore more easily excited by electromagnetic radiation. Unsaturated organic functional groups that absorb in the ultraviolet or visible regions are known as chromophores.

6. Saturated organic compounds containing such heteroatoms as oxygen, nitrogen, sulfur, or halogens have nonbonding electrons that can be excited by radiation in the 170- to 250-nm range.

7. Absorption occurs when electrons make transitions between filled and unfilled d- orbitals with energies that depend on the ligands bonded to the metal ions. The energy differences between these d- orbitals (and thus the position of the corresponding absorption maxima) depend on the position of the element in the periodic table, its oxidation state, and the nature of the ligand bonded to it.

9. Charge-transfer absorption is important for quantitative analysis because molar absorptivities are unusually large (e > 10,000 L mol 1 cm 1 ), which 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. In most charge-transfer complexes containing a metal ion, the metal serves as the electron acceptor.

10. Qualitative Applications of Ultraviolet/ Visible Spectroscopy Spectrophotometric measurements with UV radiation are useful for detecting chromophoric groups. However, ultraviolet spectra do not have sufficient fine structure to permit an analyte to be identified unambiguously. UV qualitative data must be supplemented with other physical or chemical evidence such as infrared, nuclear magnetic resonance, and mass spectra as well as solubility and melting- and boiling-point information.

11. UV spectra for qualitative analysis are usually measured using dilute solutions of the analyte. For volatile compounds, however, gas-phase spectra are often more useful than liquid-phase or solution spectra. A solvent for ultraviolet/visible spectroscopy must be transparent in the region of the spectrum where the solute absorbs. Also, the analyte must be sufficiently soluble in the solvent. Solvent polarity often influences the position of absorption maxima. For qualitative analysis, analyte spectra should thus be compared to spectra of known compounds taken in the same solvent.

12. The Effect of Slit Width Small slit widths are used for qualitative studies to preserve maximum spectral detail. Peak heights and peak separation are distorted at wider bandwidths.

13. Stray radiation occasionally causes false peaks to appear when a spectrophotometer is being operated at its wavelength extremes.

14. The important characteristics of spectrophotometric and photometric methods are: 1.Wide applicability 2.High sensitivity 3.Moderate to high selectivity 4.Good accuracy 5.Ease and convenience Molecular absorption measurements are applicable in every area requiring quantitative information: 1.Applications to absorbing species 1.Applications to Nonabsorbing species - Many nonabsorbing analytes can be determined photometrically by causing them to react with chromophoric reagents to produce products that absorb strongly in the ultraviolet and visible regions.

16. Procedural Details Wavelength Selection: To realize maximum sensitivity, spectrophotometric absorbance measurements are usually made at the wavelength of maximum absorption. Variables that influence absorption: solvent, the pH of the solution, temperature, high electrolyte concentrations, and the presence of interfering substances. The relationship between absorbance and concentration: The calibration standards should approximate as closely as possible the overall composition of the actual samples and should encompass a reasonable range of analyte concentrations. The Standard Addition Method: The composition of calibration standards should approximate the composition of the samples to be analyzed.

17. The multiple-additions method is often chosen for photometric or spectrophotometric analyses. Several increments of a standard solution are added to sample aliquots of the same size. Each solution is then diluted to a fixed volume before measuring its absorbance. When the amount of sample is limited, standard additions can be carried out by successive addition of increments of the standard to a single measured aliquot of the unknown. The measurements are made on the original solution and after each addition of standard analyte.

18. Assume that several identical aliquots V x of the unknown solution with a concen- tration cx are transferred to volumetric flasks having a volume V t. To each of these flasks is added a variable volume V s mL of a standard solution of the analyte having a known concentration c s. If the chemical system follows Beer’s law, the absorbance of the solutions is described by:

19. A plot of A s as a function of V s should yield a straight line of the form Where m is the slope and b is the intercept The unknown concentration c x can then be calculated as follows: Assuming that the uncertainties in c s, V s, and V t are negligible, the relative variance is

23. 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. Wavelengths should be selected so that the molar absorptivities of the two components differ significantly. Thus, at 1, the molar absorptivity of component M is much larger than that for component N. From the known molar absorptivities and path length, A 1 =  M1 bc M +  N1 bc N A 2 =  M2 bc M +  N2 bc N

26. The accuracy and precision of spectrophotometric analyses are often limited by the indeterminate error, or noise, associated with the instrument. The relationship between the noise encountered in the measurement of T and the resulting concentration uncertainty can be derived by writing Beer’s law in the form Taking the partial derivative, we get  c can be interpreted as the uncertainty in c Dividing the two equations, results in

28. Concentration errors When  T = k 1. For many photometers and spectrophotometers, the standard deviation in the measurement of T is constant and independent of the magnitude of T.

31. Concentration errors when  T = k 2  T 2 +  T This error originates in the shot noise that causes the output of photomultipliers and phototubes to fluctuate randomly abut a mean value. Concentration errors when  T = k 3 T The relative standard deviation in concentration from this type of uncertainty is inversely proportional to the logarithm of the transmittance. This type of uncertainty is important at low absorbances (high transmittances) but approaches zero at high absorbances.

32. Photometric and Spectrophotometric Titrations Titration Curves A photometric titration curve is a plot of absorbance (corrected for volume change) as a function of titrant volume.

33. Instrumentation Photometric titrations are usually performed with a spectrophotometer or a photometer that has been modified so that the titration vessel is held stationary in the light path. Application Photometric titrations often provide more accurate results than a direct photometric determination because the data from several measurements are used to determine the end point. The presence of other absorbing species may not interfere since only a change in absorbance is being measured.

34. The photometric end point has also been used to great advantage in titrations with EDTA and other complexing agents.

35. Spectrophotometry is a valuable tool for determining the composition of complex ions in solution and for determining their formation constants. The three most common techniques used for complex-ion studies are (1)the method of continuous variations, (2)the mole-ratio method, and (3)the slope-ratio method.

36. In the method of continuous variations, cation and ligand solutions with identical analytical concentrations are mixed such that the total volume and the total moles of reactants in each mixture are constant but the mole ratio of reactants varies systematically. The absorbance of each solution is then measured. The corrected absorbance is plotted against the volume fraction of one reactant, that is, V M /(V M + V L ), where V M is the volume of the cation solution and V L is the volume of the ligand solution.

37. In the mole-ratio method, a series of solutions is prepared in which the analytical concentration of one reactant (usually the metal ion) is held constant while that of the other is varied. A plot of absorbance versus mole ratio of the reactants is then prepared. If the formation constant is reasonably favorable, two straight lines of different slopes that intersect at a mole ratio that corresponds to the combining ratio in the complex are obtained.

39. The slope-ratio approach is particularly useful for weak complexes but is applicable only to systems in which a single complex is formed. The method assumes (1) that the complex-formation reaction can be forced to completion by a large excess of either reactant, (2) that Beer’s law is followed under these circumstances, and (3) that only the complex absorbs at the wavelength chosen.

40. 26 B Automated photometric and spectrophotometric methods Instrumentation

41. Sample and Reagent Transport System

42. Sample Injectors and Detectors Sample sizes for flow-injection analysis range from 5 to 200 mL, with 10 to 30 mL being typical for most applications. For a successful determination, it is important to inject the sample solution rapidly as a plug, or pulse, of liquid; in addition, the injections must not disturb the flow of the carrier stream. The most useful and convenient injector systems are based on sampling loops similar to those used in chromatography. Advanced Flow-Injection Techniques Flow-injection methods have been used to accomplish separations, titrations, and kinetic methods. These methods include flow reversal FIA, sequential injection FIA, and lab- on-a- valve technology.

44. 26 C Infrared absorption spectroscopy Infrared spectroscopy is a powerful tool for identifying pure organic and inorganic compounds because almost all molecular species absorb infrared radiation. However, it is a less satisfactory tool for quantitative analyses than its ultraviolet and visible counterparts because of lower sensitivity and frequent deviations from Beer’s law. Additionally, infrared absorbance measurements are considerably less precise. The energy of infrared radiation can excite vibrational and rotational transitions, but it is insufficient to excite electronic transitions.

46. Instruments for Infrared Spectrometry Three types of infrared instruments: 1.Dispersive spectrometers (spectrophotometers), 2.Fourier transform spectrometers, and 3.Filter photometers. The first two are used for obtaining complete spectra for qualitative identification, while filter photometers are designed for quantitative work. Fourier transform and filter instruments are nondispersive, that is, neither uses a grating or prism to disperse radiation into its component wavelengths.

47. Dispersive infrared instruments are similar in general design to the double- beam (in time) spectrophotometers excepting the location of the cell compartment with respect to the monochromator. Fourier transform infrared (FTIR) spectrometers offer the advantages of high sen- sitivity, resolution, and speed of data acquisition. Fourier transform instruments contain no dispersing element, and all wavelengths are detected and measured simultaneously using a Michelson interferometer. In order to separate wavelengths, it is necessary to modulate the source signal and pass it through the sample in such a way that it can be recorded as an interferogram. The interferogram is subsequently decoded by Fourier transformation, a mathematical operation that is conveniently carried out by the computer.

49. Qualitative Applications of Infrared Spectrometry An infrared absorption spectrum contains an array of sharp peaks and minima that are useful for the identification of functional groups are located in the shorter-wavelength region of the infrared.

50. Quantitative Infrared Spectrometry Quantitative infrared absorption methods differ from their ultraviolet and visible counterparts because of the greater complexity of the spectra and the narrowness of the absorption bands. The intensity of the radiation passing through the sample is compared with that of the unobstructed beam. Alternatively, a salt plate may be used as a reference. The resulting transmittance is often <100%, even in regions of the spectrum where the sample is totally transparent.

51. Applications of Quantitative Infrared Spectroscopy