Chromatography Forum

"The activated polysacchraide was desalted by gel filtration on a 1*20cm column of sephadex G-50 in 0.2 M sodium borate at PH 8.0". There, is the "0.2 M sodium borate at PH 8.0" elution solvent? I can not find the what is solution in the context.
Michelle

The gel filtration is used for buffer change. So column is equilibrated in the borate buffer and a small volume of your activated polysacharide solution is loaded on the top of the column (for desalting, no more than 1/3 of bed volume to avoid overlapping of the non retained high molecular weight analyte peak with the salt peak).
After loading of the sample, the elution is done by the same buffer that you used for equilibration and the polysaccharides will be eluted in borate buffer. It's not a binding technique.

The polysaccharides are too big to enter in the pores of that kind of column, so they elute with void volume, while low molecular weight components of the loaded solution are retained within the pores of the beads and elute later (after 1 column volume)

Hope this answer to your question

Martin

Dear Martin,
Thank you for the reply. I prefer to plus a question.
If the sample I want to separate is a simulfication solution, could these ingrediets be devided by Gel filtration? For example, there are surfactants, oil, polymers, inorganic salt in the sample. Or the micelle of surfactant and oil will be eluted together?
Michelle

Michelle,

Depends on the CMC and the size of the micelles. For exemple if you use octyl glucoside which has a high CMC and a low micelle size, there will be no problem to remove it by desalting. But, depending on its concentration, you may not be able to remove Triton X-100 (I talk by experience).
I don't know what you mean by oil, but if it's phospholipids, they can form mixed micelles with detergent and so elute together, or they can form liposomes on the column upon detergent removal (if it's an easy removable one) and elute with carbohydrates.

If your goal is to remove polymer contaminants, gel filtration could not be the best technique.

Good luck
Martin