Argentation HPLC fatty acids

The sterols include that group of compounds which contain an alcoholic hydroxyl group at C3 and a branched aliphatic chain of at least eight carbon atoms at C17. These compounds, which include cholesterol and its derivatives, can occur as free alcohols or as long-chain fatty acid esters. A number of HPLC techniques have been used in the analysis of the sterols including reversed phase, non-aque-ous reversed phase, normal phase, argentation and combinations of the above. 

Since its introduction by Morris and others in 1962, TLC on silica gel impregnated with silver nitrate has been of enormous value to the lipid analyst. It is sometimes termed "argentation" chromatography. The basis of the separation is the facility with which the double bonds in the alkyl chains of fatty acids form polar complexes reversibly with silver compounds. Fatty acids can be separated according to both the number and the configuration of their double bonds and sometimes, with care, according to the position of the double bonds in the alkyl chain. HPLC has been slow to make a mark in this area, because of problems in preparing stable columns, but many of the major difficulties now appear to have been resolved. However, most of the data on the elution characteristics of silver complexes of unsaturated fatty acids has been obtained by TLC. 

Fig. 4. Argentation HPLC of two /ra 5-C18 l fatty acids as phenacyl esters. Nucleosil lOSA-AgNOj, column length 250 mm, i.d. 4mm, Temp. -15 C. Flow rate 0-4ml/min. CHCI3 hexane, 1 13. UV detection 243nm.

In the present state of scientific knowledge, control of the amount of long-chain fatty acids in the human diet is desirable. Suitable— though not rapid, easy or inexpensive—methods of analysis have been developed for the determination of erucic acid in foods. For laboratories which have the capability of capillary GLC, this method would appear to be the best and most rapid, since no sorting test would be necessary. However, as an official reference method, the argentation TLC/GLC procedure is preferred because it requires less specialised apparatus, less experimental art, and is more easily defined in documentary form. When it is further developed, HPLC will also play a part in certain laboratories. 

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