Supercritical CO2 extraction of solid-state cultivation fungus producing azaphilone polyketides

Supercritical Fluid Extraction (SFE) methods dedicated to microorganisms specialized metabolites are very scarce in the literature and limited to liquid cultivation. We proposed here a new sample preparation method to achieve SFE of specialized metabolites from solid-state cultivation. SFE parameters, including CO2 pressure, temperature of extraction cell and percentage of co-solvent, were optimized in the case of solid-state cultures of Penicillium sclerotiorum SNB-CN111, a filamentous fungus producing azaphilone pigments. The metabolic composition of the extracts was then analyzed by reverse-phase liquid chromatography coupled to electrospray ionization and tandem mass spectrometry in data dependent acquisition mode. The resulting molecular networks generated by MetGem software allowed the annotation of the extracted metabolites in the different conditions, confirming the enrichment of fractions according to the polarity of azaphilone subfamilies. First, the 100 % CO2 fraction a yield ten times higher than hexane maceration The optimization of SFE method led to an extraction yield twice as high as ethyl acetate maceration when mixing CO2 with ethanol and, indicating that CO2/ethanol SFE is more environmentally-friendly and efficient than standard maceration methods for the extraction of azaphilones from solid-state fermentation.