C18 or C8

[lotussss]

Hello,

I have a complex mixture to separate. It's about a molecule and its synthesis impurities. Visibly there is compounds with differents functions. What conclusion can I explain? : when I use a C18 column all compounds are almost separated, except 2 which are coelued. When I use a C8 column I lost some separations but my 2 compounds are separated. These 2 compounds are elued in the end of the chromatogramm. I think there is problem of size of molecule. What do you think about that? I don't arrive to separate all my compounds with the same column and the same system. what can I do?

Thanks

[Wayne Way]

You could try a gradient or a different chemistry column such as an embedded polar group column.

[JBush]

what about trying a smaller particle size C8 column?, what have you tried in variations of these two columns?

[tom jupille]

On average, there is at least as much selectivity difference between a randomly chosen C18 column and a randomly chosen C8 column as there is between two randomly chosen C18 columns.

In reversed-phase, you have essentially six variables which you can use to change selectivity:
- solvent strength (gradient steepness)
- temperature
- solvent type
- column type
- pH
- additives.

the use of additives and pH is sometimes problematic (long equilibration time for additives, and possible robustness issues for pH). Of the other four, three of them (solvent strength, solvent type, and temperature) are continuous variables which are easily "tweaked". The fourth (column type) is essentially a discontinous variable (each column is unique).

So, if this were my problem, here's what I would do:
1. start with either your C8 or the C18 column.
2. change the gradient steepness (double or halve the gradient time) and see if the peak spacing changes. If not, go on to the next step. If it does change, spend some time to find the optimum steepness.
3. Make a big change in temperature (e.g, from 35 to 50 degrees) and see if the peak spacing changes. If not, go on to the next step. If it does change, spend some time to find the optimum temperature.
4. Switch your organic solvent from ACN to MeOH (or vice versa) and see if the peak spacing changes. If not, go on to the next step. If it does change, spend some time to find the optimum blend of ACN and MeOH.
5. Depending on how badly you dislike THF, you might try repeating step 4 with THF.
6. Try a different column. As suggested by Wayne, an EPG column would be most likely to change selectivity. Go back to Step 2 with the new column.