Course # CHEM 724

(Size Exclusion Chromatoghraphy)

By
Afshan Gul
Size-exclusion chromatography
The Apparatus
 lower limit upper limit
dextran gels
Sephadex G-50 1.5 30
Sephadex G-75 3 80
Sephadex G-100 4 150
Sephadex G-150 5 300
Sephadex G-200 5 600
polyacrylamide gels
Bio-Gel P-10 1.5 20
Bio-Gel P-30 2.5 40
Bio-Gel P-60 3 60
Bio-Gel P-100 5 100
Bio-Gel P-150 15 150
BioGel P-200 30 200
Bio-Gel P-300 60 400
dextran-polyacrylamide gels
Sephacryl S-200 5 250
Sephacryl S-300 10 1500
Sephacryl S-400 20 8000
agarose gels
Sepharose 6B 10 4000
Sepharose 4B 60 20,000
Sepharose 2B 70 40,000
Bio-Gel A-0.5 10 500
Bio-Gel A-1.5 10 1500
Bio-Gel A-5 10 5000
Bio-Gel A-15 40 15,000
Bio-Gel A-50 100 50,000
The following parameters should be accounted for in
applications of size exclusion chromatoghraphy:

Column dimensions

Flow rate.

Mobile phase

Sample volume
 Advantages of Size Exclusion Chromatography

Separations can be carried out under any appropriate

condition

Labile substances are not affected b/c of no adsorption

Less zone spreading

Simple calculation of Molecular weight

 Applications of Size Exclusion Chromatography

 Purification of enzymes and proteins

 Large scale preparation:

1. Separation of product from reactants.

2. Separation of inhibitors from enzyme preparation
3. Removal of contaminants
4. Separation of DNA from proteins
5. Purification of proteins
 Applications of Gel Chromatography

 Analytical Tool:
1. Determination of plasma proteins in diagnosis of various
diseases
2. Study of binding b/w proteins and small molecules
3. Fractionation of polymers

 Desalting And Buffer Exchange

THANKS