Apple ring rot, caused by the pathogenic fungus Botryosphaeria dothidea , has inflicted substantial economic losses and caused significant food safety concerns. In this study, a pimarane-type diterpenoid, diaporthein B (DTB), isolated from a marine-derived fungus, exhibited significant antifungal activity against B. dothidea , with an EC 50 value of 8.8 μg/mL. Transcriptome, metabolome, and physiological assays revealed that DTB may target mitochondria and disrupt the tricarboxylic acid (TCA) cycle and oxidative phosphorylation processes. This interference led to increased accumulation of reactive oxygen species and subsequent lipid peroxidation, ultimately inhibiting fungal growth. Furthermore, DTB exhibited an inhibitory potency against apple ring rot at a concentration of 31.2 μg/mL, achieving rates ranging from 67.7 to 81.6% across four distinct apple cultivars. These results indicated that DTB could serve as a novel fungicide for controlling apple ring rot in apple cultivation, transportation, and storage.