The spectrophotometric methods of analysis covered in this chapter include those based on the absorption, emission, or scattering of electromagnetic radiation. When a molecule absorbs UV/Vis radiation it undergoes a change in its valence shell electron configuration. A change in vibrational energy results from the absorption of IR radiation. Experimentally we measure the fraction of radiation transmitted, T, by the sample. Instrumentation for measuring absorption requires a source of electromagnetic radiation, a means for selecting a wavelength, and a detector for measuring transmittance. Beer’s law relates absorbance to both transmittance and to the concentration of the absorbing species (\(A = - \log T = \varepsilon b C\)).
In atomic absorption we measure the absorption of radiation by gas phase atoms. Samples are atomized using thermal energy from either a flame or a graphite furnace. Because the width of an atom’s absorption band is so narrow, the continuum sources common for molecular absorption are not used. Instead, a hollow cathode lamp provides the necessary line source of radiation. Atomic absorption suffers from a number of spectral and chemical interferences. The absorption or scattering of radiation from the sample’s matrix are important spectral interferences that are minimized by background correction. Chemical interferences include the formation of nonvolatile forms of the analyte and ionization of the analyte. The former interference is minimized by using a releasing agent or a protecting agent, and an ionization suppressor helps minimize the latter interference.
When a molecule absorbs radiation it moves from a lower energy state to a higher energy state. In returning to the lower energy state the molecule may emit radiation. This process is called photoluminescence. One form of photoluminescence is fluorescence in which the analyte emits a photon without undergoing a change in its spin state. In phosphorescence, emission occurs with a change in the analyte’s spin state. For low concentrations of analyte, both fluorescent and phosphorescent emission intensities are a linear function of the analyte’s concentration. Thermally excited atoms also emit radiation, forming the basis for atomic emission spectroscopy. Thermal excitation is achieved using either a flame or a plasma.
Spectroscopic measurements also include the scattering of light by a particulate form of the analyte. In turbidimetry, the decrease in the radiation’s transmission through the sample is measured and related to the ana- lyte’s concentration through an equation similar to Beer’s law. In nephelometry we measure the intensity of scattered radiation, which varies linearly with the analyte’s concentration.
attenuated total reflectance
fluorescent quantum yield
method of continuous variations
phosphorescent quantum yield
singlet excited state
triplet excited state