Oleanolic acid (1) is a bioactive compound widely distributed in nature. Microbial transformation is an effective way to alter the structures of compounds. The result of biotransformation of oleanolic acid was carried out in this paper. Four transform products (2-5) from 1 by Penicillium melinii were isolated. Their structures were established by spectral data interpretation as 21β-hydroxyoleanolic acid (2), 21α-hydroxyoleanolic acid (3), canthic acid (4), and 7α, 21β-dihydroxyl oleanolic acid (5). Compound 3 had stronger antibacterial activities against MRSA and Staphylococcus aureus than the substrate. It was reported for the first time that the hydroxylation products of oleanic acid can be obtained by biotransformation using incubation with Penicillium melinii.