I have to determine Fatty acids with a chainlenght between C10 and C22. I use a RP18 cloumn and a PDA-Detector. The signal of the mixture of those acids are too weak. So I also need a usefull Preperation method. Can anybody help me?

Have you considered GC after making methyl esters? That's what the soap industry does. HPLC is not always the answer to every issue.

Gregor,

The most widely applied derivatization method for the HPLC determination of fatty acids utilizes dibromoacetophenone to produce p-bromophenacyl esters which are readily detected in the UV. You should be able to find detailed information on this methodology in any good reference to derivatization in HPLC. I found the following link which may be helpful in a search on Google: http://www.cardinal.com/pts/pdf/HPLC%20Determination%20of%20Free%20Fatty%20Acids%20Roman%20et%20al%20Cardinal%20Health%20PTS.PDF It gives an example application for Saw Palmetto.

Hi Gregor, unfortunately I can only provide you with an application using two GPC columns in series, THF as mobile phase. 30 to 4 carbon fatty acids in THF are separated. But still, if possible try GC and contact the guys at Restek. I know they have an excellent application database (Varian, Agilent, Supelco and VICI also). Or you have to do derivatisation, as Chris Pohl mentioned. Good luck. Gerhard

I had success with a Water's free fatty acid column and a RI detector. Do you have to use the PDA? This was about 5 years ago and I don't know if the column is still available. The fatty acids were resolved in order of increasing carbon number. The analysis was complicated by the presence of unsaturated fatty acids. However, we were able to resolve the items in our mixture.

As an addendum to Gerhard Kratz's posting, Restek manufactures a series of columns for organic acids and carbohydrates. One I have used in the past for small molecular weight organic acids is the ROA Organic Acid Column. These are ion exclusion columns and are basically suflonated styrene divinyl benzene (and salt forms). Very simple mobile phase. You may want to contact them as Gerhard recommended to see if they would be applicable to fatty acids. However one drawback here, if you derivatize the acid functionality, you will defeat the separation mechanism of ion exclusion. GC is by far the best way to go.

It's probably worth mentioning given Bill Tate's suggestion above that ion exclusion columns are really not practical with fatty acids in the 10-22 carbon chain length range. These acids are not even soluble in water under acid conditions and so would require a very solvent rich mobile phase. High solvent content mobile phases can be used with resins of this type but standard commercial products are designed to be used with a aqueous eluents, so a custom product packed in a high solvent content mobile phase would be required to have any chance of separating this class of analytes. Assuming you don't have ready access to the GC, p-bromophenacyl esters are your best bet for analysis via HPLC.