Hydrophobic interaction chromatography (HIC) is an effective method for purification and separation of proteins (especially enzymes) based on differences in their surface hydrophobicity. Since this method does not use organic solvents like reversed phase chromatography, there is only a little loss in enzyme activity and the tercially structure of proteins (denaturation). COMSOSIL 5HIC is a packed column for hydrophobic interaction chromatography on silica gel base.

Characteristics

  • No need for organic solvents.(Protein is not denatured.)
  • Allows high pressure and high flow rate because of silica gel base.
  • No problem with swelling like with polymer gel bases.
  • Economic solution.

Application

Separation of protein standards
11
Column:COSMOSIL 5HIC 4.6mmI.D.×50mm
Mobile phaseA: 20 mmol/l Phosphate buffer
+100 mmol/l Na2SO4 + 1.5mol/l (NH4)2SO4(pH6)
B: 20 mmol/l Phosphate buffer
+100 mmol/l Na2SO4(pH6.7)
A 100 %->B 100 % 10min linear gradient
Flow rate1.0 ml/min
Temperature30 C
DetectionUV 220 nm
Sample1. Myoglobin 0.5 mg/ml
2. B-Lactoglobulin 1.0 mg/ml
3. Hemoglobin 2.5 mg/ml
4. BSA 1.0 mg/ml
Injection vol.Mixture 2.0 µl
In hydrophobic interaction chromatography, at first the high (NH4)2SO4 concentration buffer (1~2 mol/l) is used, to facilitate adsorbing of proteins to the stationary phase. Next, lower salt concentration applied gradually to remove the proteins of lower hydrophobicity from the stationary phase.

Separation of Crude B-Glucosidase

21
Column:COSMOSIL 5HIC 4.6mmI.D.×50mm
Mobile phaseA: 20 mmol/l Phosphate buffer
+100 mmol/l Na2SO4 + 2.0 mol/l (NH4)2SO4(pH6)
B: 20 mmol/l Phosphate buffer
+100 mmol/l Na2SO4(pH6.7)
A 100 %->B 100 % 10 min linear gradient
Flow rate1.0 ml/min
Temperature30 C
DetectionUV 220 nm
Sample10.0 mg/ml
Injection vol.1.5 µl
MATERIAL CHARACTERISTICS
 
COSMOSIL 5HIC
Silica gel
high purity porous spherical silica
Average particle size
5 µm
Average pore size
approx. 300Å
Specific surface area
approx. 150m²/g

Cosmosil 5HIC Column

ItemCatalogPrice ($U.S.)Buy
5HIC – Analytical Column, 4.6 x 50mm 08882-00 $412.00 Add to cart