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Abstract In a previous study, we reported that Curcuma longa strain Ryudai gold (RD) showed antifungal activity against Fusarium solani sensu lato (FSSL) among the different species and varieties of turmeric. The present study focused on isolation, identification and structural elucidation of antifungal compounds in RD. The ethyl acetate (EtOAc) fraction was eluted with n -hexane and EtOAc with gradually increasing the concentration of EtOAc (n -hexane:EtOAc; 100:0; 80:20; 60:40, 40:60, 20:80 and 0:100). The antifungal compounds were isolated from the most effective fraction by using silica gel, TOYOPEARL® HW-40F column, and high-performance liquid chromatography. Structural identification of the antifungal compounds was conducted using 1H NMR, 13C NMR, and liquid chromatography-tandem mass spectrometry. The MeOH extract of the rhizome of RD inhibited the growth of FSSL in a concentration-dependent manner. The EtOAc fraction of the MeOH extract of RD demonstrated the highest antifungal activity against FSSL. The purified antifungal compounds were turmeronol B (1), turmeronol A (2), (E)-α-atlantone (3), dihydrobisdemethoxycurcumin (4), demethoxycurcumin (5) and curcumin (6). These six compounds showed concentration-dependent antifungal activity against FSSL. The concentration required for 50% growth inhibition (IC 50) of the four isolates of FSSL ranged from 116 to172, 127 to 185, 88 to 109, 90 to 112, 74 to 80 and 63 to 68 μM/L for turmeronol B, turmeronol A, (E)-α-atlantone, dihydrobisdemethoxycurcumin, demethoxycurcumin and curcumin, respectively. These results suggested that RD contained potential antifungal compounds that could be useful to control FSSL. The isolated compounds of RD can be a good source of natural antifungal agents or the lead compounds for the development of new synthetic drugs. Graphical abstract Unlabelled Image Highlights • Ryudai gold is a strain of C. longa developed by the University of the Ryukyus and registered by the Ministry of Agriculture, Forestry and Fisheries, Japan. • We have purified 6 potential antifungal compounds namely turmeronol B (1), turmeronol A (2), (E)-α-atlantone (3), dihydrobisdemethoxycurcumin (4), demethoxycurcumin (5) and curcumin (6). • The purified compounds showed concentration-dependent antifungal activity against Fusarium solani sensu lato. [ABSTRACT FROM AUTHOR]