Z Lebensm Unters Forsch A (1999) 208 : 203–207 Q Springer-Verlag 1999 L. Cossignani 7 M. S. Simonetti 7 A. Neri 7 P. Damiani (Y) Istituto di Chimica Bromatologica, Università degli Studi, Via S. Costanzo, P. O. Box 346, I-06100 Perugia, Italy e-mail: dapi@unipg.it ORIGINAL PAPER L. Cossignani 7 M. S. Simonetti 7 A. Neri P. Damiani Structural analysis of triacylglycerol fraction and some its sub-fractions from an eicosapentaenoic acid/docosahexaenoic acid dietary supplement Received: 18 February 1998 / Revised version: 25 June 1998 Abstract Ag c - and RP-HPLC have been used to study the triacylglycerol fraction of a dietary supple- ment containing fish oil. The fatty acid (FA) distribu- tion in the glycerol backbone of the triacylglycerol frac- tion was determined by stereospecific analysis. Eicosa- pentaenoic acid, the most abundant poly unsaturated FA, was predominantly located in the sn-3 position, while docosahexaenoic and docosapentaenoic acids were generally located in the sn-2 position. The nutri- tional significance of these findings is briefly dis- cussed. Key words Triacylglycerols 7 Silver-ion and reverse-phase high-performance liquid chromatography 7 Structural analysis 7 Eicosapentaenoic acid 7 Docosahexaenoic acid Introduction The effects of the introduction of poly unsaturated fatty acids n-3 (PUFA n-3) into the human diet have been investigated [1] for several years. PUFA n-3 are modu- lators of the metabolism of arachidonic acid to eicosa- noids [2] and they are important for the normal growth of the retina and brain [3]. Many studies on hyperlipi- demic patients have shown that the consumption of PUFA n-3 protects then against cardiovascular diseases and hypertension in several ways [1, 4]. Dietary satu- rated fatty acids (FA) (especially palmitic acid) and cholesterol accelerate atherogenesis, whereas mono un- saturated FA and PUFA n-3 reduce the levels of plas- matic lipoproteins (i.e. very low-density lipoproteins and low-density lipoproteins) and cholesterol [4–6]. Many studies have demonstrated that dietary changes can be related to a rapid change in the FA composition of plasma lipids [6, 7]. Among foods, the lipid fraction of fish is rich in PUFA n-3 as eicosapentaenoic acid (EPA, C20 : 5 n-3) and docosahexaenoic acid (DHA, C22 : 6 n-3); for these reasons dietary supplements based on fish oil are commonly used. The lipid fraction of fish oil is also characterized by various FA which differ in terms of length of C chain, and number and position of double bonds. This leads to the presence of a great number of molecular species of triacylglycerols (TAG). The FA composition of TAG (% mol of total lipid fraction) of fish oil has been determined by Ag c - HPLC [8]. Some authors [9] have combined Ag c - HPLC, to obtain TAG separation according to the de- gree of unsaturation, with RP-HPLC in which the re- tention parameters of TAG molecular species, depend- ing on chainlength and/or on total number of double bonds, have been evaluated in terms of partition num- ber [10], equivalent C number [11–14] and other analo- gous algorithms such as selectivity [15]. Since dietary lipids are absorbed mainly as free FA or as sn-2 monoacylglycerols resulting from the action of lipases, the position occupied by FA in the glycerolic backbone is important for physiological and nutritional reasons; consequently, stereospecific analysis is also an important tool with which to establish the nutritional quality of TAG [16, 17]. In this work the stereospecific analysis of a fish oil dietary supplement was carried out to determine the distribution of FA (especially PUFA n-3) in the TAG glycerolic backbone. Ag c -HPLC analysis of TAG was achieved and the most abundant sub-fractions obtained were submitted to RP-HPLC to obtain the required in- formation. Materials and methods Materials. The dietary supplement was refined fish oil concen- trate. Sample preparation. The TAG fraction was isolated from the die- tary integrator by TLC (silica gel plates, 0.25 mm, 20 cm!20 cm)