HIGH-PRESSURE LIQUID-CHROMATOGRAPHY OF OXO-EICOSANOIDS DERIVED FROM ARACHIDONIC-ACID

Eicosanoids are a large group of biologically active metabolites of ar achidonic acid and related C-20 fatty acids. Many of these compounds c ontain hydroxyl groups which can be converted to oxo groups by a varie ty of substrate-specific dehydrogenases. In many cases, this results i n a reduction in potency, but in others, such as the oxidation of B-hy droxyeicosatetraenoic acid to its oxo metabolite B-oxo-eicosatetraenoi c acid, there is a dramatic increase in biological activity. Thus, it is often very important to analyze the relative amounts of oxo- and hy droxyeicosanoids formed by various cells and tissues. The present stud y was designed to compare the chromatographic behavior of oxo-eicosano ids and their hydroxy counterparts in commonly used mobile phases for reversed-phase and normal-phase HPLC. We examined three groups of eico sanoids: prostaglandins, leukotriene B-4 and some of its metabolites, and monohydroxy-eicosanoids and their oxo metabolites, We found that i n reversed-phase HPLC, the retention times of oxo-eicosanoids were lon ger than those of the corresponding hydroxy-eicosanoids in mobile phas es containing acetonitrile as the major organic component, whereas the reverse was true for mobile phases containing methanol. Normal-phase HPLC using mobile phases containing hexane, isopropanol, and acetic ac id gave excellent separation of oxo- and hydroxy-eicosanoids. Increasi ng the concentration of acetic acid in the mobile phase selectively re duced the retention times of oxo-eicosatetraenoic acids compared to mo nohydroxy-eicosatetraenoic acids, whereas the reverse was true for iso propanol. Differences in the chromatographic behavior of oxo- and hydr oxy-eicosanoids can be useful clues in the structural characterization of these compounds, as illustrated by the chromatographic properties of a complex series of LTB(4) metabolites formed by rat neutrophils. ( C) 1997 Academic Press.