I'm thinking of using either 2D paper chromatography or column chromatography. Which do you think is better if I have many compounds on my sample and I want to estimate their quantity?
In my opinion, it (roughly) depends on what kind of compounds you except to be present in your sample. Are we talking sugars? low molecular weight carboxylic acids? small phenolics? resin acids? flavonoids? lignans? etc, you get the point.
Paper chromatography is better for separation of polar to very polar compounds (due to the fact that your paper contains water and is polar). If you would do column chromatography you would check fractions on TLC, right? TLC is, compared to paper chromatography, better at separating slightly polar to nonpolar compounds. This could of course be enhanced somewhat by the eluent system, though.
I don't know if you have access to GC & GC-MS (the former for quantitative analysis, the latter for qualitative analysis), but I would perhaps use that instead. Of course this means the compounds should be volatile; you could run silylated samples to make them more volatile. And add an internal standard (IS) with a known concentration to your sample to be able to calculate the amount of different compounds (based on their peak area) in your sample. This approach requires that your IS doesn't have the same retention time as one of the compounds in your sample, though.
If GC isn't an alternative, I would perhaps go for column chromatography, but if you have many compounds it might take long/you might need to do many columns.
Thanks for your input Jan-Erik. I am interested in the hydrolyzable and condensed tannin component of my extract, so I think I will use paper chromatography because it is a cheaper alternative to your other aforementioned chromatographic techniques. I do not have access to GC in our lab and I think it will be more expensive because of the standards I require specially if I have many components in my extract. Also, I don't think tannins are volatile enough for GC.
Alright, best of luck! We analyze tannins as trimethylsilyl derivatives in our lab by GC (acid depolymerization and carbocation capture by phloroglucinol, which yields the monomers and phloroglucinol adducts).
Do you have a published protocol for that? Maybe I can also use it if I'm able to use a GC here in our university.
Hernes, P.J; Hedges, J.I., Anal. Chem. 72, 2000, 5115-5124. It's been used for whole samples, though.
You could also look into The Tannin Handbook by Ann E. Hagerman. It might give some fruitful information (e.g., the acid butanol assay for condensed tannins).I hope it helps.
http://www.users.muohio.edu/hagermae/
Thanks a lot! That is a very informative site. I hope I can put all of it into good use.
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