Hi Everyone!

I have a very polar guanidine which is not retained (ret. vol ~5-6 ml)very well on a C18 column even using a weak solvent like 95:5 0.1%TFA:methanol. Can someone please suggest other systems? Is ion exchange chromatography a better solution? I would like the retention volume to be about 15 ml. thanks

ganesan

You can do several things. One suggestion is to use a reversed-phase column that can be used in a solvent with a higher water content. Examples are either an unendcapped C18 column (Resolve C18) or one of these new phases with an incorporated polar groups (SymmetryShield RP18). I would go the second route, since with the older unendcapped C18 you would have to fight for getting a good peak shape.
The alternative is to explore hydrophilic interaction chromatography (HILIC). You can use a silica column as the stationary phase. HILIC works the other way round than reversed-phase chromatography: no retention in water and more retention in acetonitrile/buffer. I would start with around 75% acetonitrile with a buffer. If the retention is too low, increase the acetonitrile content. If it is too low, reduce it.

I think your chances are much better by cation exchange chromatography. Since a compound of this type is very strongly basic, the separation can probably be readily accomplished over a wide pH range on either a silica or a polymer based cation exchange column.

Hi Chris,
I don't see the difficulty. He is getting a retention factor of around 1 to 1.5 with 5% methanol. Taking out the methanol should get him in exactly the right spot....
Uwe

We succussesfully reteined the aminoguanidine on polymeric column with 100% water (pH 11.0).
Dr.A.Weisman, Chemagis Ltd., Israel