Comparative Metabolomics of Fungal Strains

Three hyphal mass discs (7 mm) of each strain were incubated in PD broth for 7 days at 28 °C and 180 rpm, followed by mass collection through vacuum filtration [44 (link)]. The biomass was dried, weighed (WT, 3.14 g; ΔAosec22, 2.89 g), and used to prepare extracts of the same concentration. Ethyl acetate (250 mL) was added to extract the fermentation broth and ultrasonic extraction was performed for 40 min. The fermentation broth was allowed to stand for 12 h and the crude extract was concentrated in ethyl acetate under vacuum [44 (link)]. The crude extract was dissolved in 500 µL of analytic-grade methanol and the solution was filtered through a 0.22 µm membrane filter for liquid chromatography–mass spectrometry (LC–MS) analysis. The metabolic profiles of the WT and mutant strains were compared using the Thermo Xcalibur software (Thermo Fisher Scientific). Untargeted metabolomics was performed using Compounds Discoverer 3.0 software (Thermo Fisher Scientific) [49 (link)].