Chromatography Forum

I am developing a method to measure the conversion of tall oil fatty acid to a dimer. I have been using a stabiliwax column from restek. The column was purchased new, conditioned following their directions, and the baseline looks great. The difficulty I'm running into is that I am unable to use derivitized samples, because the column has a polyethelene glycol phase.

At this point the best peak shape and best separation has been an isothermal method.

Initial temp: 240C
Final Hold: 30 min

While the separtaion has vastly improved over the first few methods I was working with there is still quite a bit of coelution around where I expect to see the C16-C18 peaks.

Would it be easier to derivitize, dry and then return my sample to solution or have others obtained resonable results from underivitized fatty acids on PEG columns?

Another method that has provided resonable separation, less co-elution, but awful peak shape is:

Initial Temp: 120C
Initial hold: 2min
Ramp: 15deg/min
Final Temp: 240C
Final hold: 30min

All of my samples have used either DMF or Methylene chloride as a solvent. MCl seems to provided slightly clearer results on the carbowax column. Usually our derivitizing agent has been 1% TMCS. Much of this analysis was previously (and currently) done with derivitized samples on a restek Rtx-50 column.

I don't have a lot of experience in this area myself but I can tell you what's working for me...

I have used Restek FAMEWAX and Supelco Omegawax for complicated mixtures of derivatized saturated and unsaturated fatty acid samples and they both work great with oven gradients. I am using iso-octane + 0.05 g/L BHT as the diluent (the BHT is there on the hopes that it will prtect the unsaturated components from oxidation!), with methyl tricosanoate internal standard. I haven't deliberately tried to analyze free fatty acids (FFAs) with these particular methods, but there is a recent thread in the forum where some people have suggested that this will not work well, and I have circumstantial evidence to back that up.

If you want to analyze FFAs, both Phenomenex and HP that I'm aware of have columns designed for FFA analysys, the ZB-FFAP and HP-FFAP respectively... I think the Restek Stabiliwax DA is equivalent... while from the sounds of it trying to do FFA analysis directly is bound to be unpleasant and should be avoided if you can, maybe you could give one of these a whirl? If you do, let me know how it works out!

Be ready to clean out your inlets a lot and watch the split vent filter, depending on the samples it can gum up surprisingly fast!

Thanks much for the suggestions! I'll let you know where things end up for me on this.