Ultraviolet-Visible (UV) Spectroscopy- Definition, Principle, Uses

U.V spectroscopy involves the promotion of electrons (n, σ, π) from the ground state to a higher energy state. In U.V. spectroscopy, the excitation of the electron occurs in the range of 200-800 nm.

Ultraviolet-Visible (UV) Spectroscopy
Ultraviolet-Visible (UV) Spectroscopy

Principle of Ultraviolet-Visible (UV) Spectroscopy

When the electromagnetic radiation of the correct frequency passes through the compound’s sample, some amount of energy is absorbed by the sample, which causes the excitation of an electron from the lower energy level to the higher energy level. The radiation on leaving the sample after absorption will be either less intense, or its intensity may be completely lost.

The absorption of light by a molecule is given by Beer-Lambert law as:

A= log Io/I = εc l

A= absorbance

Io = intensity of incident light

I= intensity of transmitted light

ε= molar excitation coefficient

c= concentration of solution in mole/cm-1

l= pathlength of sample

Types of Electronic Transitions

According to the molecular orbital theory, when a molecule is excited by energy absorption, the transition of electrons occurs from bonding to antibonding orbitals.

The various electronic transition is σ-σ*, n-σ*, π-π*, n-π*. The energy required for various transition obey the order of σ-σ*> n-σ*> π-π*> n-π*.

σ-σ* transition= It is a high-energy process. Saturated hydrocarbon like methane shows σ-σ* transition.

n-σ* transition = Saturated compounds containing one heteroatom show n-σ* transition. Here the transition from nonbonding orbital to sigma antibonding orbital takes place. This type of transition is sensitive to hydrogen bonding. Hydrogen bonding shifts absorption towards a lower wavelength.

CH3Cl= 172-175 mμ.

CH3I= 258 mμ. ( this is due to the low electronegativity of iodine)

π-π* transition= Molecules having double and triple bonds show π-π* transition. This type of electronic transition normally occurs at lower energy.

e.g. Ethylene = 175 nm

n-π* transition = In this transition, one electron of the heteroatom gets excited to π antibonding orbital. This process requires low energy.

n-π*= 290nm,

π-π*= 180 nm

Selection rules

The various electronic transitions are governed by different restrictions called selection rules. They are:

  • Transitions involving a change in the spin quantum number of an electron during the transition, do not occur.
  • Transition between the orbital of different symmetry does not occur. e.g., n-π* transition is symmetry forbidden.

The intensity of absorption is directly proportional to the transitional probability. An allowed transition will have an εmax value greater than 1000.

Different terms used in Ultraviolet-Visible (UV) Spectroscopy

  • Chromophores: Chromophores are the compounds that are responsible for imparting color to the compounds. They absorb the light in the U.V. region. E.g. nitro group present in the nitro compounds are the chromophore which imparts a yellow color to these compounds.
  • Auxochromes: Auxochromes are colour-enhancing group. This group doen’t itself act as the chromophore but their presence shifts the absorption band towards the longer wavelength. e.g., -OH, -OR, -NH2 etc.
  • Fluorescence: Fluorescence is the process by which a molecule emits light of a longer wavelength after absorbing different light of a short wavelength. The fluorescence process stops when the irradiating light is removed.
  • Phosphorescence: The phosphorescence involves the continuous emission of radiation of longer wavelength when the irradiating light is removed.

Absorption and Intensity Shifts

Bathochromic effect: It is an effect by which the absorption maximum is shifted towards a longer wavelength due to the presence of an autochrome or by a change of solvent. Bathochromic shifts are also called red shifts.
Hypsochromic shifts: This is the effect by which the absorption maximum is shifted towards a shorter wavelength. It is also called blue shift. It may be caused by the removal of conjugation and changing the solvent’s polarity.
Hyperchromic effect: It is an effect due to which the intensity of absorption maximum increases Emax increases. B-band for pyridine at 275 mμ, Emax 2750 is shifted to 262mμ, Emax 3560 for 2-methyl pyridine. The introduction of autochrome usually increases the intensity of absorption.
Hypochromic effect: It is defined as an effect due to which the intensity of absorption maximum decreases, i.e., extinction coefficient Emax decreases. The introduction of the group which distorts the geometry of the molecule causes the hypochromic effect.

Types of absorption bands

  • K- bands= K- bands originate due to π-π* transition from a compound containing a conjugated system. Such types of bands arise in compounds like dienes, polyenes, and enones.
  • R- bands= Such types of bands originate due to the n-π* transition of a single chromophoric group and have at least one lone pair of electrons on the hetero atom. R-bands are also called forbidden bands.
  • B-bands= This type of band arises due to π-π* transition in aromatic or hetero-aromatic molecules.
  • E-bands= Such bonds originate due to the electronic transitions in the benzenoid system of three ethylenic bonds, which are in closed cyclic conjugation.

Applications of Ultraviolet-Visible (UV) Spectroscopy

  • Detection of functional groups: U.V. spectroscopy is used to detect the presence and absence of chromophores. The absence of a band at a particular wavelength indicates the absence of a particular group in this compound.
  • Extent of conjugation: The extent of conjugation in polyenes can be estimated by using U.V spectroscopy methods.

Conjugated and nonconjugated compounds can be distinguished by using U.V methods.

  • Identification of unknown compounds: Unknown compounds can be identified by comparing their spectra with the spectra of known compounds. If two spectra coincide, these compounds must be identical otherwise different.
  • Determination of configuration of geometrical isomers: Cis-alkenes absorb at different wavelengths as compared to their corresponding trans isomers. Cis and Trans isomers can be distinguished by using U.V spectroscopy.e.g.,

Cis-stillbene λmax = 283 mμ.

Trans-stillbene λmax= 295.5 mμ.

  • Identification of compounds in different solvents: Sometimes, the structure of the compound changes with a change in a solvent which can be determined by using U.V spectroscopy.

Chloral hydrate shows an absorption maximum at 290mμ in hexane, while in aqueous solution, the absorption disappears. This is due to the presence of the carbonyl group in hexane solution as its structure is present as CCl3.CHO.H2O, whereas in an aqueous solution, it is present as CCl3.CH(OH)2.

  • It is also used in the pharmaceutical analysis process and bacterial culturing.

References

  1. Silverstein, R. M., Webster, F. X., & Kiemle, D. J. (2005). Spectrometric identification of organic compounds. Hoboken, NJ: John Wiley & Sons.
  2. https://byjus.com/chemistry/uv-vis-spectroscopy/
  3. https://byjus.com/chemistry/principle-of-uv-visible-spectroscopy/
  4. https://www.indiastudychannel.com/resources/146681-Principle-working-and-applications-of-UV-spectroscopy.aspx
  5. https://www.pharmatutor.org/pharma-analysis/analytical-aspects-of-uv-visible-spectroscopy/applications.html
  6. https://microbenotes.com/uv-spectroscopy-principle-instrumentation-applications/
  7. https://www.technologynetworks.com/analysis/articles/uv-vis-spectroscopy-principle-strengths-and-limitations-and-applications-349865

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