Chromatography Forum

Uwe,
I have been researching your suggested approach all morning, but have once again come up empty handed for similar applications. I guess my real question is this: with any of these methodologies, we are not really concerned so much with the sugar portion of the structure as we are with the anionic sulfate groups . . . is that correct?

Bryan,

b). Mobile phase conditions. Did you also try the TEA acetate?
This is volatile & compatible with ELSD

Looks like this buffer will result in a pH around 7.0. As I stated, I've worked over the entire range (even 7.0) but have not seen a difference in resolution. Is there something about THIS buffer that might effect the specificity of the separation?

c). SAX - I don't think this is a good option. The reason is because
you're limited as far as what mobile phase you can use (volatile).
I'm sure SAX will separate these compounds -
but you'll need a different detector.

When you say you are SURE SAX will separate these compounds, what makes you say that? I did attempt a separation on a PRP-X100 which resulted in the same co-elution. I DO have conductivity and RI detectors at my disposal, so if that helps ... I just don't have any experience with the conductivity detector, so I would have to figure that out too. Same for the ligand-exchange option, I guess.

I will send you some chromatograms shortly, but I don't really think they will be much help. Thanks for your advice.

dkollmorgen,

if you send me samples we will try to develop a method for you free of charge. We just did three methods for our customers last week after they struggle for few months, so there is a good chance that we can do the same for you. If you need CDA you can download one from our website:
http://www.sielc.com/Services_MethodDevelopment.html

Please let me know if you are interested.

here is example for something similar (?) - heparin:
http://www.sielc.com/compound_236.html

I believe the TEA acetate can also have some ion-pairing with the solutes.
The trick is to hit the 'sweet spot' between normal phase and anion exchange.

Since you do not require ELS or MS detection, you may want to research some
of the other column vendors (SAX, ligand exchange, ion exclusion, ect.).

It should be possible to play with ion-exclusion with the KS 802 or 803 column. I would first consider the KS 803. Manipulate the salt concentration to change the retention pattern.