Determination of molecular weight by Light scattering method.
Due to big size macromolecules in solution, turbidity is produced. The light rays get scattered, when pass through macromolecule solution due to turbidity. The turbidity T is given by
Where I₀ is the intensity of incident light.
Iₜ is the intensity of transmitted light after passing through a solution of length l.
In case of proteins or higher polymers the turbidity is small and is determine from the intensity of light scattered at 90° to the beam. This can be done by using a simple photometer as shown in figure below.
Turbidity increase with increase in concentration as well as molecular weight. The Turbidity is related to molecular weight by the equation given by Debye.
Hc/T=1/M+2Bc
Where B is second virial coefficient and H is a constant.
If we plot a graph between Hc/T vs c we get a straight line with intercept 1/M. Thus
Iₜ is the intensity of transmitted light after passing through a solution of length l.
In case of proteins or higher polymers the turbidity is small and is determine from the intensity of light scattered at 90° to the beam. This can be done by using a simple photometer as shown in figure below.
Hc/T=1/M+2Bc
Where B is second virial coefficient and H is a constant.
If we plot a graph between Hc/T vs c we get a straight line with intercept 1/M. Thus
This curve was plotted by Zimm, so is called Zimm's Curve. Thus the curve is a straight line which is extrapolated to zero concentration gives the intercept equal to 1/M can be calculated.
Limitation
1) This method gives wrong results at molecular weight M above 10,000,00.
2) Dust particles also scattered light , hence it causes problem and gives wrong result. Next
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