A novel styrylpyrone derivative, named phelliribsin B (1), as well as four biogenetically related known compounds, phellifuropyranone A (2), inoscavin C (3), inoscavin A (4), and inoscavin D (5) were separated and purified from the medicinal fungus Phellinus ribis. The structure of phelliribsin B was determined by spectroscopic analysis, and the absolute configuration was assigned by experimental and calculated ECD data. Additionally, the plausible biosynthetic pathway of 1 was also proposed. Compound 1 showed moderately cytotoxic activity against HepG2 and SKOV-3 tumor cell lines with IC50 values of 32.71 and 57.89 μM, respectively. Based on the results of cytotoxicity against HepG2 tumor cells, the structure-activity relationship of compounds 1-4 with similar skeletons was discussed. The styrylpyrone derivatives with similar skeletons have moderately cytotoxic activity and have the potential to play an important role in the anti-tumor treatment.