A major problem in the discovery of new biologically active compounds from natural products is the re-isolation of known compounds. To address this problem, dereplication strategies are needed that enable crude extracts to be screened for the presence of known compounds before isolation efforts are initiated. The specific attributes of this dereplication methodology include a focus on bioactive secondary metabolites from fungi, the use of a 10 min chromatographic method, and the inclusion of both HRMS and MS/MS data.