The instrument used to measure the amount of radiation absorbed by an organic molecule is called a spectrophotometer.
Working- Radiation from the source passes through the vertical entrance slit S1 onto the spherical mirror M1, from which a parallel beam is reflected onto the grating. The source emits white light i.e., it radiates energy over a wide range of frequencies. Interference takes place at the grid of the grating, therefore different frequencies reflect from the grating at different angles. Thus when the grating is rotated as shown, a succession of frequencies is swept across the mirror M2, from where it is focussed on to the exit slit, S2. So the frequency of the radiation arriving at S2 depends on the angle of the grating.
From S2 the radiation falls on M3, which focuses it on to the detector. The latter, responds with an electrical signal proportional to the intensity of the radiation falling on it. This signal is amplified electronically and then used either to drive a pen, when records the spectrum immediately or is collected and stored on a computer for later processing and display.
The sample can be placed almost anywhere in the radiation beam. In order to be able to minimize its size, it is often placed close to one of the slits, where the beam is smallest. It is preferable to put it near the exit slit, S2, where it is subjected to only a narrow range of frequencies at any one time, rather than near S1 where it is subjected to the full output of the source and may suffer degradation from the intense heat or light.
Components of a Spectrophotometer:
The different components of a spectrophotometer are-
(1) Source of Electromagnetic Radiation- Different sources are used in different regions, they are as-
- For ultra-violet region hydrogen discharge lamp is used as source of radiation.
- For visible region tungsten filament lamp is used as source of radiation.
- For infrared region the source of radiation is Nernst filament, which consist of oxides of cerium and thorium, which are kept at high temperature (or 1500°C).
- In microwave region Klystron or Magnetron is a source of radiation. Klystron is specially developed electron tube in which the electrons on their way towards the plate generate electromagnetic radiations of suitable frequency.
- For nuclear magnetic resonance, radio frequency radiations are used which are generated by causing electric currents to oscillate in coils of wire.
(2) Monochromator- The radiation is separated into its constituent frequencies by dispersing devices such as prism or grating. The prism used-
- For ultra violet radiation is quartz prism.
- For visible radiation is glass.
- For infrared it is made of rock salt.
(3) Sample Holder- It is used to hold the sample in the instrument. It can be-
- Test tube in colorimetry.
- Quartz cell in U.V. and visible spectroscopy.
- Two infrared transparent sodium chloride windows are used to hold the sample in IR spectroscopy.
- A glass tube called NMR tube is used in NMR spectroscopy.
(4) Detector- A detector convert the incident radiation into an electrical signal for appropriate signal processing or plotting. The photomultiplier is widely used in the optical and ultraviolet regions. A thermocouple detector is used in the infrared region. It consists of blackened gold foil to which thermoelectric alloys are attached. These alloys generate an electric current where the temperature is increased.
(5) Amplification and Recording Unit- The radiation from the detector is chopped by a shutter that rotates in a beam so that an alternating signal is obtained from the detector. This is because an oscillating signal is easy to amplify. Amplification is followed by output generation either on a computer monitor or graphical point-out.