Antifungal activities of natural substrances from Eucalyptus darlympleana, E. globules, E. gunnii and E. unigera were evaluated against postharvest pathogens of kiwifruits, Botrytis cinerea, Botryosphaeria dothidea, and Diaporthe actinidiae, to screen effective natural substances as an alternative to chemical fungicides. Methanol extract of the Eucalyptus trees showed strong antagonistic activity against the pathogenic fungi. Among them, E. unigera and E. darlympleana effectively inhibited mycelial growth of the pathogens. For chemical identification of the antifungal substances, the methanol extract of E. darlympleana leaves was successively partitioned with CH2Cl2, EtOAc, n-BuOH and H2O. Among the fractions, CH2Cl2 and n-BuOH showed strong inhibitory activity of mycelial growth of the fungi. Five compounds were isolated from EtOAc and n-BuOH fractions subjected to SiO2 column chromatography. Two phenolic compounds (gallic acid and 3,4-dihydroxybenzoic acid) and three flavonoid compounds (quercetin, quercetin-3-O-α-L-rhamnoside, quercetin-3-O-β- D-glucoside) were identified by 1H-NMR and 13C-NMR spectroscopy. Among them, only gallic acid was found to be effective in mycelial growth and spore germination of B. cinerea at relatively high concentrations. The results suggest that gallic acid can be a safer and more acceptable alternative to current synthetic fungicides controlling soft rot decay of kiwifruit during postharvest storage.