In the course of our investigations of natural products from microorganisms for drug discovery, we studied Streptomyces lydicus (strain HKI0343)by physico-chemical analysis of its metabolite pattern. Four new secondary metabolites, lydiamycins A–D (1–4; PBB = piperazic acid, PSA = 2-pentylsuccinic acid), were discovered in the fermention broth of this strain by thin-layer chromatography (visualization by UV absorption and staining with anisaldehyde/H2SO4). The cyclodepsipeptides lydiamycins A (20 mg), B (15 mg), C (10 mg), and D (7 mg)were obtained as white solids by standard chromatographic procedures from a 300-L fermentation of the strain. Herein we report the structure determination of each metabolite using a combination of CID ESIMS/MS data (CID = collision-induced dissociation)and 1D and 2D NMR techniques such as DEPT, COSY, HMBC, TOCSY, and ROESY, the assignment of the absolute configurations, and, finally, the biological properties.