Recently published 16-membered diene macrolide antibiotics including bafilomycins, hygrolidins, hygrolidincyclopentanedion amides, leucanicidin, and L-681,110 share a similar aglycon partial structure and exhibit bactericidal activity against a broad spectrum of Gram-positive bacteria as well as fungicidal and antiparasitic effects. New 16-membered macrolides without cyclic deoxy carbohydrate moieties were isolated from cultures of Streptomyces griseus Tii 2599 (DSM 2610, producer of bafilomycins A-C) and Streptomyces hygroscopicus MA5285 (ATCC 31955, producer of hygrolidin cyclopentanedion amide). This communication presents the production and isolation conditions, physical data, and structures of bafilomycin D (I), bafilomycin E (II), and oxohygrolidin (III). Bafilomycin D (I) and oxohygrolidin (III) were obtained by alkaline hydrolysis of bafilomycin Bx, bafilomycin Cl, or hygrolidin, while bafilomycin E (II) was isolated from the culture filtrate and could not be derived from hydrolysis of bafilomycin Bl under alkaline or acidic conditions. Their physical-chemical properties (molecular formula, melting point, spectral data such as NMR and MS) are described. The biological activities of I-III (differing from bafilomycins A-C and hygrolidins by lacking the cyclic deoxy pyranose partial structure) are summarized: they inhibited the growth of Gram-positive bacteria at concentrations below 100 μg/ml, showed no inhibitory effects against Gram-negative bacteria at 1000 μg/ml, exhibited fungicidal effects against Phythium ultimum, Mycosphaerella musae, and Fusarium nivale at 5 μg/ml, and had insecticidal activities against Plutella maculipennis, Phaedon cochleariae, Dysdercus intermedius, and Ceratitis capitata when applied at 1 mg/ml.