UV Visible spectroscopy (Instrumentation, working and Applications)

This video describes the instrumentation and working of UV-Visible spectroscopy. It obeys Beer-Lambert law. The complete instrumentation includes source, monochromator, beam splitter, sample holder, detector and recorder and the function of eachcomponent. This video describes the double beam spectrophotometer techniques. It discusses the applications of this technique. Ultraviolet-visible spectroscopy or ultraviolet-visible spectrophotometry (UV-Vis or UV/Vis) refers to absorption spectroscopy or reflectance spectroscopy.
UV-Visible Spectroscopy
(Instrumentation and working)
You will be able to
explain the instrumentation and working of UV-Visible spectroscopy.
list out the applications of UV-Visible spectroscopy.
compare UV-Visible spectroscopy with colorimetric analysis
When UV or visible light passes through the compound with multiple bonds, the transition of valence electrons from lower to higher energy level occurs and gives rise to UV-Visible spectroscopy.
Types of transitions in organic compounds
Source - Supplies light in the UV range (l = 200-400nm) - hydrogen or deuterium lamp and visible range (l = 400-800nm) - tungsten lamp.
Monochromator - Allows only the chosen wavelength and absorbs all the other wavelengths and this happens for the complete chosen range of wavelength. eg. prism, diffraction grating etc.
Beam splitter - Splits the beam of light of each wavelength into two halves of equal intensities, one goes to the test sample and other to the blank/reference sample.
Pair of identical cells (cuvettes) - One cell is filled with the test sample and the other with the blank sample (solvent). eg. quartz.
Detector - It detects the intensities of the transmitted light coming out of the cells and compares for each wavelength and generates current proportional to the difference in the intensities of the sample and blank. photocell, photodiode.
Recorder- It records an UV/Visible spectrograph with absorbance against the wavelength.
Note: The instrument scans in a similar manner for the complete chosen wave length range (200-400nm ).
If the sample does not absorb light at a particular wavelength, I = Io , A=0
If the sample absorb light at a particular wavelength,
Applications of UV spectroscopy
Detection of functional group - presence or absence of chromophore.
Can determine the extent of conjugation in polyenes (more the unsaturation, more the shift towards longer wavelength).
Identification of unknown compound.
Elucidate the structure of vitamin A and K (characteristic peaks).
Preference over the tautomeric forms.
Distinguish between the geometric isomers (cis and trans forms absorbs different wavelengths).
Distinguish between conformations (equitorial and axial form absorbs different wavelengths).