Three new naphtho-g-pyrones, 5-hydroxy-6,8-dimethoxy-2-hydroxymethyl-4H-naphtho [2,3-b]pyran-4-one (1, nigerasperone A), 3,3Ј-dihydro-2,2Ј,5,5Ј-tetrahydroxy-8,8Ј,10,10Ј-tetramethoxy-2,2Ј-dimethyl-(6Ј,9-bi-4H-naphtho[1,2-b]pyran)-4,4Ј-dione (2, nigerasperone B), and 3Ј-hydro-2Ј,5,5Ј,8-tetrahydroxy-6,6Ј,8Ј-trimethoxy-2,2Ј-dimethyl-(7,10Ј-bi-4H-naphtho[2,3-b]pyran)-4,4Ј-dione (3, nigerasperone C), together with nine related known compounds were characterized from Aspergillus niger EN-13, an endophytic fungus isolated from the marine brown alga Colpomenia sinuosa. Their structures were elucidated by detailed analysis of spectroscopic data and by comparison with literature reports. In the cytotoxic assay, these compounds did not show remarkable inhibitory effects against A549 and SMMC-7721 tumor cell lines. However, 3 and several known compounds showed weak antifungal activity against Candida albicans and moderate activity on DPPH scavenging.