Objective To investigate the secondary metabolites of the fungus Phialemonium sp. associated with the South China Sea cucumber Holothuria nobilis Selenka. Methods The EtOAc extract of Phialemonium sp. was purified by repeated column chromatography on silica gel, Sephadex LH-20, and high-performance liquid chromatography (HPLC). Structural determination of the isolated compounds was achieved by spectroscopic analysis and by comparison with previously reported data. Results Two depsidones were isolated and their structures were determined as: emeguisin A and aspergillusidone C. Conclusion This is the first report on the secondary metabolites of Phialemonium sp. associated with the sea cucumber H. nobilis Selenka, and the two depsidones are reported for the first time from the fungus strain Phialemonium sp.