Rezex ROA organic acid question

[primate]

Hello

I am working on quantification of galacturonic acid with this column. The samples also contains some monosaccharides : glucose, rhamnose, arabinose, xylose, galactose, mannose, fucose.

The detector is a ELSD. I got only 6 peaks in various conditions (flowrate, temperature) with these 8 compounds.

Is it due to the kind of column (maybe it has not a good selectivity for monsaccharides) or has somebody an idea to separate each compound?

Thank you for your answer.

[tom jupille]

All of those are available as pure compounds. First step would be to run standards individually and see what coelutes (or doesn't elute, as the case may be).

[primate]

The mannose, galactose and xylose are coeluted

[tom jupille]

The Resex ROA is a sulfonated polystyrene packing in the hydrogen form. The same basic packing is also available in Ca, Ag, and Pb forms, each of which give different selectivity for carbohydrates (it's been a while since I worked with them, and I don't remember which one would be specifically applicable). Check with BioRad or with Dionex; they should be able to make a specific recommendation (web site info is in the FAQ; drill down to the "additional information" section).

[rhaefe]

I am no expert in carbohydrate analysis but from the little I know it will be impossible to separate all compounds on a single column (I am referring to the classic ligand exchange approach).
Switching, lets say to a lead form will resolve mannose, galactose and xylose but arabinose, fucose and mannose will most likely co-elute.
You can check out:
http://chromatography.transgenomic.com/ ... tChart.asp

That is a pretty useful chart. If you have more questions I can give you the contact info of the person in charge there. My e-mail is:
rhaefele@hamiltoncompany.com

Maybe someone from Dionex can shed some light on the capabilities of anion exchange in NaOH with a PAD...

cheers,

Robert Haefele
(NOT affiliated with TBIO)

[Uwe Neue]

Here is another idea: since the columns with Ca or Pb give a completely different selectivity, you could use a second column to resolve the ones that are unresolved on your H column. You will need to work out, what is best - just two different columns in series, or a column switching protocol, where the unresolved peaks from the H column are parked on say a Pb column and eluted from that after the analysis form the H column is complete.

It's a bit more complicated, but if you like to tinker, this could work.

[malaka50]

Here is another idea: since the columns with Ca or Pb give a completely different selectivity, you could use a second column to resolve the ones that are unresolved on your H column. You will need to work out, what is best - just two different columns in series, or a column switching protocol, where the unresolved peaks from the H column are parked on say a Pb column and eluted from that after the analysis form the H column is complete.

It's a bit more complicated, but if you like to tinker, this could work.

i agree. add another column in series. either a rezex RCM or RPM would give the best separation. using only the ROA is not enough to separate all of the compounds.