beta-lapachone (1) has entered phases I and II clinical trials for the treatment of solid tumors and the therapeutic efficacy of beta-lapachone is closely related to its metabolic process. In order to contribute to a better understanding of human metabolism of beta-lapachone, Cunninghamella elegans ATCC 10028b was used as a microbial model of mammalian metabolism to biotransform beta-lapachone and two new glycosylated derivatives were produced. The chemical structures were elucidated as 6-hydroxy-2,2-dimethyl-3,4-dihydro-2H-naphtho[1,2-b]pyran-5-O-beta-D-glucopyranoside (2) and 5-hydroxy-2,2-dimethyl-3,4-dihydro-2H-naphtho[1,2-b]pyran-6-O-beta-D-glucopyranoside (3) by H-1 NMR, C-13 NMR, HMBC, HMQC, COSY and HRMS analyses. The major derivative (3) displayed a lower activity against breast cancer cell line SKBR-3 (IC50 = 312.5 mu M) than beta-lapachone (IC50 = 5.6 mu M), but did not show cytotoxicity against normal fibroblasts cell line GM07492-A, whereas beta-lapachone was highly toxic (IC50 = 7.25 mu M). These metabolites were reported here for the first time and are similar to those that occur in phase II of human metabolism (C) 2013 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.