DETERMINATION OF TOTAL LONG-CHAIN FATTY-ACIDS IN HUMAN PLASMA AND LIPOPROTEINS, BEFORE AND DURING COPPER-STIMULATED OXIDATION, BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

An improved method for the high-performance liquid chromatographic det ermination (HPLC) of total or free long-chain (>C12) fatty acids in sm all volumes (10 mu L) of human plasma and lipoprotein samples is descr ibed, The method is based on the formation of 2-nitrophenylhydrazine ( 2-NPH) derivatives and offers an alternative to gas chromatographic (G C) fatty acid determination, The retention of 2-NPH fatty acid derivat ives on the HPLC system differs from the typical pattern produced by G C separation, thus offering a powerful tool for confirmation of peak i dentification where GC peak resolution is poor, Fatty acids determined include saturates [myristic acid, C14:0; palmitic acid, C16:0; steari c acid, C18:0; eicosanoic acid, C20:0; docosanoic acid, C22:0; and tet racosanoic acid, C24:0], monounsaturates [palmitoleic, C16:1; petrosel enic, C18:1(n12); oleic, C18:1(n9); and erucic, C22:1], and polyunsatu rates [linoleic, C18:2; linolenic, C18:3(n3); gamma-linolenic acid, C1 8:3(n6); eicosatrienoic, C20:3; arachidonic, C20:4; eicosapentanoic, C 20:5; docosahexanoic, C20:6; and docosatetraenoic, C22:4], Mean recove ries of fatty acids added to LDL samples were 94.1 - 109.4%, and intra assay coefficients of variation for the major fatty acids in human pla sma were 2.7 - 6.9%, The potential of the method for further developme nt is discussed, Long-chain fatty acid profiles are given for plasma a nd very low-, low-, and high-density lipoprotein (before and during co pper-stimulated oxidation) from human blood.