I am trying to separate 9-cis retinoic acid and all-trans retinoic acid in a resolution solution.
When I inject them separately, 9-cis RA comes at 10.88 min. and all trans RA at 11.01 min. I have tried adjusting the mobil phase ratios and using a longer column but had no success. Any help will be greatly appreciated.

Thanks in advance

Probably you column and mobile phase combination is not selective enough.Try to change both or one of them.

Good luck,Dr.Alex Weisman
Chemagis Ltd. ISRAEL

Use different solvents. Maybe a dash of THF?

There are 2 references you might look at for the
separation of the 9, 13 and all trans isomers of
retinoic acid. Both are in J. Chromatography B.
The first, an isocratic RP system, is Disdier,
Beatrice et al, 683, p143, (1996) and the second,
a normal phase gradient system, can be found in
757,2 365-368, (2001).

I've recently separated some 5 diasteromers using reversed phase chromatography on a Develosil C30, 250 x 4.6mm, 5um column. I've been able to achieve separation with 0.1% TFA in ACN/H2O mix and also with 0.2% H2PO4 in ACN/KH2PO4 in H2O.

Had some success with Hypercarb, although you should start with 100% organic and decrease your organic strength as needed.

Pure reverse phase HPLC is not good at seperating position isomers. I believe the first seperation you got is based on the secondary stationary phase characteristics (eg Si-OH). Even if you have good resolution at the beginning, it's likely that you will not get a repeatable resolution when you use a different batch of the same column. Try it.

I will suggest you use size-exclusion chromatographic columns (SEC)

By what mechanism would SEC separate the isomers of retinol from one another? I don't see it...Have you done it? Please advise.

Best,

Chris

I have over the course of many years worked w Vit
A acid, alcohol, aldehyde, palmitate and acetate.
We separated the all trans from 9,11,13,11/13 cis
as well as the epoxide and retro forms using
either normal phase or reverse phase
chromatography. I believe if you look up the two
references above you’ll be ok. With regard to the
normal phase reference- the authors separate the
isomers of several derivatives of VA acid as well
as the isomers of the acid. You ought to be able
to pick out the conditions applicable to an
isocratic run of just the acid isomers.

Regarding SEC – My first guess would be that this
approach to be unlikely to provide a separation.
The problem here is the separation of isomers the
size of which should be equivalent. Are there in
fact references to the separation of isomers by
SEC?