Hi,

I am searching for a method for separation of alpha and beta carotene with a very good selectivity because I will use this method in future also for semipreparative scale.

Thank you for your help.

I would try normal phase (silica, diol and NH2) or C30.

I would try reversed phase. With a polymeric C18 or C30 (where polymeric refers to the bonding chemistry-silica is still the support)

Thanks for answers.

in literature I found a C30 method but the selektivity is not good enough.

on normal phase please give a example for chromatographic conditions. the carotenes are hydrophobic substances! why do you think normal phase must work?

Try using silica gel containing silver. Silver is known to bind to double bonds and the system is very sensitive to geometry of the molecule in respect to double bonds. Good luck!

I guess the next question is "how good" does the selectivity have to be? With a C30 column I can separate the geometric isomers of alpha and beta carotene, which is probably not what you want to do if you are trying to isolate one from the other. With a polymeric C18 column I get a good separation of the 2 compounds, but not of the isomers. I cannot recall the exact mobile phase composition I used for this separation, but I belive it was an acetone/water isocratic method with a fairly high concentration of acetone. It is a little counterintuitive to use acetone as a mobile phase component, but with visible detection it works fine.

I think a selectivity higher than 1,5 will be good enough if the peak form is ok. If there is a tailing, than the selectivity must be higher than 2.
To work with acetone at 450 nm can be a good idea. It is a strong solvent for such hydrophobic compounds. I will try it.

If your trying to sep c/t isomers preparatively I
would try a silica column and a hex/alcohol (IPA
or EtOH) mobile phase. I have used silver
impregnated si cols successfully w Vitamin A
however there was column bleed (Ag) and this might
be a problem w isolated fractions and the
longevity of the column.