Piericidins are insecticidal compounds isolated from mycelia of Streptomyces mobaraensis and Streptomyces pactum. They are toxic to several species of insects, aphids, and mites. Piericidin A has been shown to block electron transport between NADH dehydrogenase and coenzyme Q. The structures of the piericidin group have been elucidated, with members including piericidins An, Bn, Cn and Dn (n=l, 2, 3 and 4). In the course of our screening program to find inhibitors of phosphatidylinositol turnover, we have isolated a novel antibiotic, piericidin B1 TV-oxide, from Streptomyces strain MJ288-OF3. For production, the strain was inoculated into seed medium (sucrose 4.0%, soybean meal 2.5%, NaCl 0.25%, CaCO3 0.32%, CuSO4·5H2O 0.0005%, MnCl2·4H2O 0.0005%, ZnSO4·7H2O 0.0005%, pH 7.4) and incubated for 3 days at 28°C on a rotary shaker (180rpm), then 2ml of culture was transferred to fermentation medium (same composition as seed) and fermented for 4 days under the same conditions. Morphological and physiological studies revealed the strain resembled Streptomyces aburaviensis. The 6-liter fermentation broth was filtered, mycelia extracted with acetone, the extract combined with filtrate and extracted with EtOAc. The EtOAc extract was concentrated to an oily matter, mixed with silica gel and applied to a silica gel column, washed with CHCl3 and eluted with CHCl3-MeOH (10:1). The residue was partitioned via centrifugal partition chromatography (CPC) using CHCl3-MeOH-H2O (5:6:4) (lower phase stationary, partition coefficient 0.11). Active fractions were chromatographed on Sephadex LH-20 with EtOAc, and crude material further purified by preparative HPLC (Nucleosil 5C18 column, 80% MeOH), yielding 53mg of piericidin B1 TV-oxide. Piericidin B1 TV-oxide is a pale yellow oil soluble in MeOH, EtOAc, CHCl3 but insoluble in water. UV spectra showed maxima at 226 (ε 34,130), 238sh (ε 29,680), 246sh (ε 20,780), 267nm (ε 7,570) in MeOH; 212 (ε 35,020), 238 (ε 24,960), 246sh (ε 20,780), 275nm (ε 5,030) in 0.1N HCl-MeOH; 238sh (ε 33,240), 246sh (ε 22,300), 276nm (ε 11,130) in 0.1N NaOH-MeOH. IR spectrum (CHCl3) had absorptions at 3530, 3000, 2950, 2890 (sh), 2850, 1620, 1520, 1480, 1470, 1380, 1360, 1310, 1290, 1200, 1170, 1130, 1110, 1080, 1020, 980, 930, 880, 840cm⁻¹. Molecular formula C26H39NO5 was assigned based on HRFAB mass spectrum (m/z 446.2897 (M+H)⁺) and ¹H/¹³C NMR spectra. [α]D²⁵ was -4.5° (c 0.2, MeOH). Structure was determined by comparing ¹H/¹³C NMR with piericidin B1, mass spectral analysis (one more oxygen than B1, presumed N-oxide), chemical reduction to piericidin B1 (confirmed by FAB-MS and ¹H NMR), and optical rotation (similar to B1, assigning C-9/C-10 configurations as S-S), concluding the structure as piericidin B1 TV-oxide. Phosphatidylinositol turnover was assayed via EGF-stimulated [³H]inositol incorporation into A431 cell phospholipids. Piericidin B1 TV-oxide inhibited turnover with IC50 1.2μg/ml (stronger than B1's IC50 5.0μg/ml). Antimicrobial activities: active against Gram-positive/negative bacteria and fungi, while B1 was not. Thus, piericidin B1 TV-oxide is a new piericidin family member with both antibacterial and phosphatidylinositol turnover inhibitory activities.