UV spectroscopy principle and instrumentation :- by sarfaraz khan

1. UV spectroscopy
Sarfaraz Khan
Assistant Professor
Quality Assurance

9. Frequency and wavelength are inversely proportional

15. Electronic Transitions
When a molecule absorbs Ultra-violet or Visible light, its electrons get
promoted from ground state to the higher excited state results in electronic
transitions.

25. Verification of Beer lambert law
From Beer Lambert’s law, we have
A=εCl
This law can be verified by plotting the absorbance(A) against the molar
concentration (C) of the solution. A straight line passing through the origin
is obtained.

26. Deviations from Beers law
When we plot absorbance against a straight line passing through the origin
is obtained.
But a plot of absorbance and concentration deviates from linear behavior.

30. C. Chemical deviation :
Association, dissociation, or ionization of solutes causes deviation
Presence of some other substance that absorbs at the same wavelength
as the solute may cause deviation.
The presence of some impurities in the colored compounds may cause
deviation

42. Instruments for measuring the absorption of U.V. or visible
radiation are made up of the following components;
1. Sources (UV and visible)
2. Filter or monochromator
3. Sample cells
4. Detector
INSTRUMENTATION

43. Components of spectrophotometer

49. Range of wavelength 200-400 nm

55. Components of monochromatic

71. Sample cells:
• The cell holding the sample should be transparent to the wavelength
region to be recorded.
• Quartz or fused silica cuvettes are required for spectroscopy in the UV
region.
• The thickness of the cell is generally 1 cm. cells may be rectangular in
shape or cylindrical with flat ends.
• Cuvette from Quartz can be used in UV as well as in visible
spectroscopy.
• Cuvette from Glass is suitable for visible but not for UV spectroscopy
because it absorbs UV radiation.

74. Photovoltaic cell
Photovoltaic cell is also known as barrier layer or photronic cell.
It consists of a metallic base plate like iron or aluminium which acts as one
electrode.
On its surface, a thin layer of a semiconductor metal like selenium is
deposited.
Then the surface of selenium is covered by a very thin layer of silver or gold
which acts as a second collector tube.

75. When the radiation is incident upon the surface of selenium, electrons are
generated at the selenium- silver surface and the electrons are collected by the
silver.
This accumulation at the silver surface creates an electric voltage difference
between two electrodes.
When it is connected to a galvanometer, a flow of current is observed which is
proportional to intensity and wavelength of light falling on it.

76. Phototubes
Phototubes are also known as photo emissive cells.
A phototube consists of an evacuated glass bulb.
There is light sensitive cathode inside it.
The inner surface of cathode is coated with light sensitive layer such as
potassium oxide and silver oxide.
When radiation is incident upon a cathode, photoelectrons are emitted.
These are collected by an anode causing current to flow.
Current is amplified and recorded.
More sensitive as compared to barrier layer cell therefore widely used.

78. Photomultiplier tube
It is a commonly used detector in UV spectroscopy.
It consists of a photo emissive cathode (a cathode which emits electrons when
struck by photons of radiation), several dynodes (which emit several electrons
for each electron striking them) and an anode.
A photon of radiation entering the tube strikes the cathode, causing the
emission of several electrons.
These electrons are accelerated towards the first dynode (which is 90V more
positive than the cathode).
The electrons strike the first dynode, causing the emission of several
electrons for each incident electron.
These electrons are then accelerated towards the second dynode, to produce
more electrons which are accelerated towards dynode three and so on.

79. The electrons are collected at the anode.
The resulting current is amplified and measured. Photomultipliers are
very sensitive to UV and visible radiation.
They have fast response times.
It is extremely sensitive to light and is best suited where weaker or low
radiation is received.