BIOSYNTHESIS OF ARACHIDONIC-ACID METABOLITES IN LIMULUS-POLYPHEMUS AMEBOCYTES - ANALYSIS BY LIQUID CHROMATOGRAPHY-ELECTROSPRAY IONIZATION MASS-SPECTROMETRY
Jc. Macpherson et al., BIOSYNTHESIS OF ARACHIDONIC-ACID METABOLITES IN LIMULUS-POLYPHEMUS AMEBOCYTES - ANALYSIS BY LIQUID CHROMATOGRAPHY-ELECTROSPRAY IONIZATION MASS-SPECTROMETRY, Biochimica et biophysica acta, L. Lipids and lipid metabolism, 1303(2), 1996, pp. 127-136
Eicosanoid metabolites were generated by isolated granular amebocytes
of the primitive arthropod, Limulus polyphemus, when stimulated by the
calcium ionophore A23187 and/or exogenous arachidonic acid. The metab
olites were isolated, identified, and the major metabolite was quantif
ied using reverse-phase high pressure liquid chromatography (RP-HPLC)
coupled to electrospray ionization mass spectrometry (ESI-MS). Qualita
tive examination revealed putative metabolites and the major product,
8-hydroxyeicosatetraenoic acid (8-HETE), which was quantified using st
andard curves generated from extracted ion profiles of the molecular i
on. Electrospray ionization of the HETEs in negative ion mode produces
a base peak for all isomers which corresponded to the molecular ion [
(M-H)(-): m/z 319]. The molecular ion was accompanied by the neutral l
oss of water and carbon dioxide [(M-H -H2O)(-): m/z 301; (M-H -H2O-CO2
)(-): m/z 257], as well as daughter ions which were dependent upon the
position of hydroxy substitution. Standard curves were generated in f
ull scan mode for standards ranging from 6.25 to 100 ng, whereas selec
ted ion recording was used for the lower levels of 0.8 to 6.25 ng.