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Principles of Colorimetry Explained

Colorimetry is a technique for quantitatively estimating colors using a colorimeter, which measures light absorption in colored solutions. It is based on Beer’s Law and Lambert’s Law, which relate light absorption to the concentration and path length of the absorbing substance. Applications include detecting substance concentrations, impurities, and characterizing organic compounds.

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123 views2 pages

Principles of Colorimetry Explained

Colorimetry is a technique for quantitatively estimating colors using a colorimeter, which measures light absorption in colored solutions. It is based on Beer’s Law and Lambert’s Law, which relate light absorption to the concentration and path length of the absorbing substance. Applications include detecting substance concentrations, impurities, and characterizing organic compounds.

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Colorimetry

Photometers measure intensity of absorbed, transmitted or reflected light. Absorption


of light is related to color intensity Intensity of colour is proportional to concentration
of the chemical substance responsible for producing the color.

Colorimetry is the technique that uses quantitative estimation of colours. Colorimeter


is the device used to measure the amount of light absorbed by a coloured solution.
Colorimetry can be used if the substance to be analyzed is coloured, or if it can be
made coloured by a chemical reaction. Coloured solutions have the property of
absorbing light of definite wavelength

Parts of a colorimeter

[Link] source: usually a filamentous bulb


[Link]: Used for selecting monochromatic light. The filter will absorb light of
unwantedwavelength and allow only monochromatic light to pass through. This is
usually thecomplementary color.
[Link] holder called cuvette
[Link] or photo cell: It converts the radiant energy into an equivalent amount of
electricalenergy
[Link] displayer

Principle
Colorimetry is governed by 2 laws.
1. BEER'S LAW:
The amount of light absorbed by a colored solution is directly proportional to
the concentration of the absorbing substance in the solution.
AαC

2. LAMBERT’S LAW: The amount of light absorbed by a colored solution is


directly proportional to the depth of the solution (path length) through which light
passes.
If ‘L’ is the depth,
A αL

Combining the two laws


A α C α L
A= K XCXL

If
A1 - Absorbance of test solution
A2 - Absorbance of standard
C1 - Concentration of test solution
C2 - Concentration of standard

C1 = A1
C2 A2

C1 = A1 x C2
A2

= Absorbance of test x concentration of standard


Absorbance of standard

Amount of substance present per 100 ml of serum sample =

Absorbance of test x concentration of standard x 100 ml


Absorbance of standard volume of serum

Procedure
Suitable reagents are added to the substance to be analyzed to develop a color.
Intensity of color is measured and compared with a standard (known concentration of
substance)

Applications of Beer-Lambert’s Law in colorimetry & spectrophotometry:


•Detection of concentration of substances
•Detection of impurities
•Structure elucidation of organic compounds
•Monitoring dissolved oxygen content in freshwater and marine ecosystems
•Characterization of proteins
•Detection of functional groups
•Respiratory gas analysis in hospitals
•Molecular weight determination of compounds
•The visible and UV spectrophotometer may be used to identify classes of
compounds in boththe pure state and in biological preparations

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