UV

The energies of photons in the ultraviolet and visible region of the spectrum correspond to the separation of electronic energy levels in molecules and atoms. Absorption in the visible/ultraviolet can occur if two criteria are met:

• The product of Planck's constant and frequency must correspond to the difference in energy of the ground electronic state and an electronic excited state, often with an increment of a few multiples of a vibrational energy, because the electronic excitation may be accompanied by vibrational excitation.
  • In dilute solution (3 mg/l) in cyclohexane, benzene shows absorption in the range 240-280 nm (roughly 40,000 cm^-1), G Fig. 1.5. This absorption band contains several (perhaps 4, depending on the details of the measurement) subbands (known as "fine structure"), that are equally spaced in energy, with a spacing on the order of 1000 cm^-1. The fine structure arises from the resolution of transitions to different vibrational excited states, accompanying the electronic excitation common to all bands in the vibrational fine structure.
  • Benzene also shows another, more intense band at higher energy (lower wavelength), that arises from a different electronic transition.
  • Atactic polystyrene exhibits a uv absorption spectrum similar to that of benzene, or toluene, although the resolution of the vibrational fine structure may not be quite as strong.
  • A chromophore is that part of the molecule responsible for the absorption under discussion. Benzene, toluene, and polystryene all have the same chromophore, which is the six-membered aromatic ring.
  • Vibrational fine structure is most easily resolved for isolated chromophores in vacuo. It tends to become blurred in condensed states, and particularly so when the chromophore is in a strongly interacting solvent.

• The electronic transition must be accompanied by a nonzero electronic transition moment (a quantum mechanical construction that depends on the wavefunctions for the electron in its initial ground state and in its final electronic state). The extinction coefficient in Beer's Law depends on the size of the electronic transition moment.
  • In general, the extinction coefficient is small for transitions of nonbonding electrons to excited pi states. This type of transition occurs in molecules with carbonyl groups (polyesters, polyamides, etc.)
  • The extinction coefficients tend to be larger for transitions of pi electrons to excited pi states. This type of transition occurs in unsaturated hydrocarbons and also in polyesters, polyamides, etc.
  • For polyamides, which exhibit both types of transitions, the n-pi* transition occurs at lower energy (high wavelength), but it is sufficiently close to the pi-pi* transition so that it may be completely swamped (and not resolvable) by the latter transition in a common absorption measurement using unpolarized light. But it may become easily resolvable if the measurement uses polarized light, and if the sample is chiral, as in a circular dichroism spectrum of a chiral poly(amino acid) in the conformation known as the alpha helix.

Quantitative analysis of solutions

Ultraviolet absorption is often used to determine the concentration (via Beer's Law) of dilute solutions of polymers that contain chromophores. This use requires knowledge of the value of the extinction coefficient for that polymer in the solvent being used for the measurement. It can also be used to monitor kinetic processes in which a chromophore is created or destroyed, or in which a chromophore changes state. For example, ionization of a phenol, as in poly(4-vinylphenol), is accompanied by a red shift of the absorption and a strong increase in the value of the extinction coefficient.

Quantitative analysis of conformational change

Ordered arrays of identical (or similar) chromophores often have different values of the extinction coefficient than do random arrays of the same chromophores.

• The stacking of the aromatic rings in the DNA double helix is accompanied by a reduction in the values of their extinction coefficient. This phenomenon is called hypochromism. Therefore the thermally induced disruption of the DNA double helix can be monitored at 260 nm, by looking for the temperature at which there is a dramatic increase in the intensity of the absorption, due to the randomization of the arrang of chromophores.
• In contrast, the different stacking of the amide units in the alpha helix of polypeptides produces an increase in their extinction coefficient. This phenomenon is called hyperchromism. Therefore thermal disruption of the alpha helix of poly(L-alanine) is accompanied by a decrease in the intensitiy of absorption around 200 nm.
• The relative orientation of the electronic transitions moments in the ordered array is responsible for the alteration in the extinction coefficient. The transition moments are oriented differently in the DNA double helix and in the alpha helix.

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