Hi,
I would like to know if how do you pack a C18 stationary phase in an ordianary glass column and if what is the solvent system that you use in isolating compounds.
thank you very much,
Ivan,
It is the same as with normal phase silica but you use polar solvents like aqueous methanol or aqueous ACN. For example, you can use MeOH/Water mixtures. If you want to use gradient elution you can start with the appropriate aqueous methanol mixture and then use increasing amounts of MeOH.
Hope this will help.
It is better to pack with MeOH and then equilibriate with the required water/MeOH mixture. (You can increase the ratio of water stepwise).
Do you have RP TLC plates? If you do, you can test how the compunds you are interested move on RP. This will help you to determine the MeOH/Water mixtures that you use to elute the column. (Same as with normal phase column but for gradient elution you start with a high polar solvent and decrase the polarity)
yes i do have rp tlc. can you give me sample ratio of water and methanol?
I usually start with MeOH/H2O ratio of 8:2. If the compound(s) do not move or the Rf values are too low then I increase the MeOH. If the Rf values are too high you can increase the amount of H2O.
(Using a solvent system which gives a Rf value between 0.2 and 0.3 for a particular compound when you do the column chromatography will elute the compound in a reasonable time)
You can also use ACN instead of MeOH.
Also use MeOH/EtOAc mixtures if you need a slovent mixture of lower polarity than MeOH.
Regarding the Column chromatography, you can wash the column with EtOAc to clean it up at the end as it will elute the more non-polar compounds that do not elute with aqueous MeOH or MeOH.
Hope this is helpful.
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