Countercurrent chromatographic isolation of α'-propyl-substituted furan fatty acid ethyl esters from an eicosapentaenoic acid ethyl ester oil

Furan fatty acids (FuFAs) are lipophilic antioxidants that occur in small amounts (≤1% of the total fatty acids) in plants and animals. They are characterised by a furan substructure within an otherwise saturated acyl chain. Although FuFAs are valuable dietary components due to their beneficial effects, research is hampered by the unavailability of standard substances such as α'-propyl-substituted FuFAs, which are particularly common in fish oils. Here we succeeded in isolating 11-(3,4-dimethyl-5-propylfuran-2-yl)-undecanoic acid ethyl ester (11D3-EE) from gram-amounts of an eicosapentaenoic acid ethyl ester (EPA-EE) oil. After an initial enrichment by hydrogenation and urea complexation, the concentrated FuFA-EE fraction was separated by countercurrent chromatography (CCC) in two-dimensional heart-cut mode using the biphasic solvent system n- hexane/benzotrifluoride/acetonitrile (20:7:13, v / v / v ). Comparative investigations showed that the head-to-tail mode was better suited than the tail-to-head mode. With the help of the ProMISE2 software, two time-shifted injections were carried out in one run with in total four heart-cuts. As a result, 1.2 mg of 11D3-EE with a purity of 95% (recovery: ∼2.5%), 11.3 mg 11D3-EE with a purity of 90% (recovery: ∼23.6%), and 1.2 mg of of the trace compound 9D3-EE (95% purity; recovery not determinable) were isolated from the sample initially enriched by hydrogenation and urea complexation. • Isolation of two furan fatty acids (FuFAs) from an ethylated fish oil fraction. • Combining hydrogenation, urea complexation and countercurrent chromatography (CCC). • Application of two-dimensional heart-cut CCC to overcome co-elutions of FuFA-EE. • Simulation of suitable conditions for isolating FuFA-EE with ProMISE2 software. • Final purity of up to 95% of the rare α'-propyl-substituted 11D3-EE and 9D3-EE.