Chromatography Forum

Please help
I am already in Thailand to do my Diplomarwork (origninally from Europe). Therefore I have all my papers in Austria and no englisch or German Book to look
I want to seperate antioxidative Compounds of an methanolic plant extract (propably Phenolic Substances, especially terpenoids and Saponins )by column chromatography. I have only the possibility to use Normalphase(Silica - gel) or perhaps Sephadex ... what are the most useful mobile phases???
Should I change the phase (gradient).
Please answer me quickly....
Thank you so much
pharmaceutical Student

Dear Aquilegia:

I am unable to find any application in my database for such stuff by normal phase.

Anyway, typical mobile phase for that are mixes of hexane and isopropanol, dioxane and THF, just to name a few.

I would start with several isocratic injections decreasing hexane (90:10, 75:25, 50:50, 25:75) and see what happens...

I hope this helps.

Aquilegia,

In my far-off days working in synthetic carbohydrate chemistry, for analysis of reaction mixtures I used

-----silica gel TLC plates (i.e. normal phase) with,

(A) ethyl acetate/hexane (3:1) for reactions that had blocked all (or most) of the -OH groups

(B) ethyl acetate/ethanol/water (10:3:2) or (45:5:3) for those mixtures that contained many unsubstituted -OH groups.

Multiple development (2-4 times) before visualization gave an indication of the chromatography to be expected on an open column.

I hope that this helps.

JMB

Use silica, not Sephadex. Run a gradient from hexane to isopropanol. What detection will you use? If you have the capability to do low UV, you may be able to see most of the things that you are interested in.

I have decided to make first an separation of my compounds by the use of different polar and non polar solvents because I have very polar Substances in my plant extract an others (expected to be terpenoids and Saponins) which are quite unpolar. I think it woul be different to seperate them in a coloumn.
Now I have got a little Problem because of some compound which make saponification. Not easy to work with or to seperate.
We can use here UV and I think the Saponins have UV aktivity.
Just to see seperation by solvent I use TLC and anisaldehyde-sulfuric-acid als detection (spray-reagent).
Later on I search my fractions for antioxidant activity by DPPH assay.
Here is only the possibility to use normale phase which will not be easy I think because some compounds are very polar... therefore the thought with sephadex.
Thank you all very much for answering me... good help!!!!!