The Spectrophotometry

By Ain & Tini

What is spectrophotomery?
• Spectrophotometry is a
technique used to
measure how much
radiant energy a
substance absorbs at
varying wavelengths of
light.
 Spectrophotometer..
• the combination of two devices: a
spectrometer and a photometer
• The most physical lab equipments
• Spectrometer-for producing light of any
selected wavelength or color
• Photometer-for measuring the intensity of
light
 Types of spectrophotometry
• Ultaviolet(UV), visible light, Infrared(IR) microwave
• UV spectrophotometry particularly useful in
detecting and quantifying colorless substances in
solution
• IR spectrophotometry is used mostly to study the
molecular structures of complex organic compounds
• In astronomy, spectrophotometry studies also
include X-ray and gamma ray range of the spectrum
 How it works?
• The two devices are placed at either side of cuvette filled with a
liquid.
• Spectrometer produces light of desired wavelength and it passes
through the tube and reach the photometer that measures its
intensity
• Then the photometer produces a voltage signal to display device,
usually galvanometer
• As the amount of light absorbed by the liquid changes,the signals
also changes
• The concentration of a substance in solution can be measured by
calculating the amount of absorption of light at the appropriate
wavelength or a particular color
 example...
• Pigment such as chlorophyll and other colored
materials (ie,dyes)absorb energy in the visible
light range (380-760nm) while other
substances absorb at shorter wavelengths
(ultraviolet=UV) or at longer wavelengths
(infrared=IR)
• By measuring the absorption of a substance, all
the substance wavelengths which it absorb, it is
possible to identify it or at least place it in a
particular class of compounds.
• The absorption maximum is very useful when
trying to identify an unknown
• By creating and measuring a series of standards,
it is possible to quantify the amount or
concentration of a substance in a sample.
• For example:-the nucleic acid can be quantified
by absorption measurements in the UV range
Demo..

spec2_medium.wmv