help! for: RP-HPLC of hexanic lyophilized crude extract ?

[nir]

I intend to separate on RP-HPLC hexanic lyophilized crude extract of skin and mucus. The components of the crude are unknown but it is very hydrophobic.
I am very new in the field of chromatography and would be grateful for help with the following questions:
1. Which substance should I used for dissolving the sample
2. Which column recommended for this task
thanks
Nir

[Jack Silver]

Extracted in Hexane?

This suggests a very non-polar mixture.

Some questions that would help us to better help you?
-What have you tried, and what were the results?

-Why are you starting with reverse phase? Very non-polar compounds are often successfully purified with silica.

-How much material are you purifying? Is this prep scale or are you determining the presence/concentration of a compound? If analysis, are you using a published procedure?

-For reverse phase, water is the weak solvent. Methanol, acetonitrile, or other solvents are the strong solvent that let your compound elute. For a compound extracted in hexane, I would try THF as a solvent (less polar than the others).

For silica based C18 one other trick I may try, if the compound doesn't elute with THF, is non-aqueous reverse phase. Methanol takes the place of water as the polar solvent, any less polar solvents (such as dichloromethane or acetone) can be used to elute the compound- don't do this with polymeric reverse phase. For some examples of this technique with a flash chromatography system, please see: http://www.isco.com/WebProductFiles/App ... ld_C18.pdf