Chromatography Forum

Currently I am working on carotenoids. I am trying to separate carotenes and xanthophylls by SPE method (C18 and C18-OH) . I used hexane as washing solvent and acetone as eluting solvent. however i could not be able to separate them totally. Do any one suggest me any better method !!!!

Separate them from a matrix (if so what?) or from each other? Depending on which separation, how you are not able to separate them totally may need additional clarification. Give more details about what you observe.

What are you putting on the cartridge - and in what solvent?

I am using commercially available tablets for Optimizing the HPLC-MS-MS method. But in future I will work with real samples. I want to separate carotene's from xanthophylls by using SPE. Im using hexane and methanol as conditioning and equilibrating solvents, while hexane as wash solvent and acetone as eluting solvent. when i run aectone fraction I still observe the carotene fraction in it.

I assume the carotenes wash off in the hexane fraction? If so, what happens if you wash with more hexane before attempting to exlute the xanthophyls?

I read where the column is being washed with methanol and hexane. As these solvents are immiscible, it may affect your chromatography. I couldn't read the order in which the solvents are being used, so this may not be an issue. If you are eluting in the order methanol->acetone->hexane, it won't be an issue.

You may consider using dichloromethane instead of acetone (elute methanol->dichloromethane->hexane).

Here's a small amount of work I've done in this area:
http://www.isco.com/WebProductFiles/App ... idants.pdf

Hopefully, this may give you other ideas.