Chemical fungicides are an important component in disease management for most crops. As part of a program to discover natural product-based fungicides, several sensitive assay systems have been developed for the evaluation of naturally occurring antifungal agents. In this study, we focused on the discovery and evaluation of new natural product-based fungicides from Angelica sinensis (Dong Quai) for use in control of strawberry anthracnose caused by Colletotrichum species. A. sinensis oil was evaluated for antifungal activity against three Colletotrichum species using direct-bioautography assay and a strawberry detached leaf assay using C. fragariae. Chemical composition and fingerprinting were conducted by gas chromatography and gas chromatography-mass spectrometry. Major components were identified as (Z)-ligustilide (43.1%), (E)-3-butylidene phthalide (14.5%), (Z)-beta-ocimene (12.9%), and apiole (11.2%).