The entomopathogenic bacteria Xenorhabdus/Photorhabdus inhibit insect immune responses by inhibiting eicosanoid biosynthesis. Especially, the bacterial secondary metabolites inhibit PLA2 that release eicosanoid biosynthesis precursors. Some organic extracts of the bacteria-cultured broth possessed PLA2 inhibitory activity. This study used butanol to extract the metabolites and showed that the extract possessed potent inhibitory activity against insect immunity. The inhibition was reversed by the addition of eicosanoid biosynthesis precursor, suggesting PLA2 inhibition. Furthermore, sub-fractionation of the butanol extract separated the secondary metabolites and the fractions exhibited differential PLA2 inhibition. The active fractions appeared to contain novel compounds that are not known in PLA2 inhibition of these bacteria.