Ws. Powell et al., HIGH-PRESSURE LIQUID-CHROMATOGRAPHY OF OXO-EICOSANOIDS DERIVED FROM ARACHIDONIC-ACID, Analytical biochemistry, 247(1), 1997, pp. 17-24
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.