Simple Summary Insect immune responses defend fatal attacks from entomopathogens. A Gram-negative Xenorhabdus hominickii exhibits high entomopathogenicity against lepidopteran insects. During the pathogenic processes, the bacteria suppress host insect immune responses by inhibiting phospholipase A2 (PLA2) enzyme activity with the bacterial secondary metabolites. PLA2 catalyzes eicosanoid biosynthesis. Eicosanoids mediate both cellular and humoral immune responses against various insect pathogens. This study identified eight different PLA2 inhibitors from the bacterial culture broth. Butanol extract of the culture broth possessed high potency to inhibit PLA2 activity. Subsequent fractionations led to identification of eight different compounds. The synthetic compounds also showed PLA2 inhibition and insecticidal activities. Furthermore, the addition of the bacterial PLA2 inhibitors significantly enhance other bacterial pathogenicity, suggesting its potential to be applied for developing novel insecticides. Abstract Eicosanoids mediate both cellular and humoral immune responses in insects. Phospholipase A2 (PLA2) catalyzes the first committed step in eicosanoid biosynthesis. It is a common pathogenic target of two entomopathogenic bacteria, Xenorhabdus and Photorhabdus . The objective of this study was to identify novel PLA2 inhibitors from X. hominickii and determine their immunosuppressive activities. To identify novel PLA2 inhibitors, stepwise fractionation of X. hominickii culture broth and subsequent enzyme assays were performed. Eight purified fractions of bacterial metabolites were obtained. Gas chromatography and mass spectrometry (GC-MS) analysis predicted that the main components in these eight fractions were 2-cyanobenzoic acid, dibutylamine, 2-ethyl 1-hexanol, phthalimide (PM), dioctyl terephthalate, docosane, bis (2-ethylhexyl) phthalate, and 3-ethoxy-4-methoxyphenol (EMP). Their synthetic compounds inhibited the activity of PLA2 in hemocytes of a lepidopteran insect, Spodoptera exigua , in a dose-dependent manner. They also showed significant inhibitory activities against immune responses such as prophenoloxidase activation and hemocytic nodulation of S. exigua larvae, with PM and EMP exhibiting the most potent inhibitory activities. These immunosuppressive activities were specific through PLA2 inhibition because an addition of arachidonic acid, a catalytic product of PLA2, significantly rescued such suppressed immune responses. The two most potent compounds (PM and EMP) showed significant insecticidal activities after oral administration. When the compounds were mixed with Bacillus thuringiensis (Bt), they markedly increased Bt pathogenicity. This study identified eight PLA2 inhibitors from bacterial metabolites of X. hominickii and demonstrated their potential as novel insecticides.